Organization of thoracic sympathetic influences on renal nerve activity
afferent
LYNNE C. WEAVER, LINDA J. MACKLEM, KEITH A. REIMANN, ROBERT L. MECKLER, AND ROBERT S. OEHL Department of Physiology, Michigan State University, East Lansing, Michigan
LYNNE C., LINDA J. MACKLEM, KEITH A. REIROBERT L. MECKLER, AND ROBERT S. OEHL. Organization of thoracic sympathetic afferent influences on renal nerve activity. Am. J. Physiol. 237(l): H44-H50, 1979 or Am. J. Physiol.: Heart Circ. Physiol. 6(l): H44-H50, 1979.-The specific influences of thoracic sympathetic afferent nerves on sympathetic outflow to the kidney have not been defined clearly, although preliminary experiments have shown that electrical stimulation of these afferent nerves causes excitation of multifiber renal nerve activity followed by inhibition of discharge. The organization of this complex reflex was investigated further in anesthetized, vagotomized, sinoaortic-denervated cats. The apparent ability of the cardiopulmonary afferent nerves to excite and then inhibit multifiber renal nerve activity initially was attributed to opposite responses of individual renal neurons to afferent stimulation. This hypothesis was not confirmed in the present study because many single postganglionic renal fibers responded to electrical afferent stimulation with excitation followed by inhibition of discharge. Alternatively, the possibility was considered that different thoracic sympathetic afferent neurons have opposite influences on renal nerve activity. In support of this suggestion, sympathetic afferent fibers activated by stretch of the ventricles or aorta caused reflex excitation of renal nerve activity whereas activation of afferent fibers from the pulmonary vasculature caused an inhibition of this discharge. Thus, the complexity of thoracic sympathetic afferent influences on sympathetic outflow to the kidney is likely related to heterogeneity within the afferent rather than efferent neural population. WEAVER,
MANN,
electrophysiological
analysis; reflex organization;
cat
nervescanhavecomplex influences on sympathetic outflow to the kidney. Activation of these afferent fibers by a brief electrical stimulus has been shown to evoke an excitatory burst in renal nerve activity followed by inhibition of discharge (25). The neural organization responsible for the biphasic character of this response is not understood. This response can be produced by activation of a homogeneous group of myelinated A6 fibers. Because the response was observed in multifiber recording of renal nerve activity, it may reflect heterogeneity within the renal efferent population. That is, activation of a group of similar sympathetic afferent fibers may excite some renal efferent neurons while inhibiting spontaneous activity in others. Sympathetic nerves innervate several vascular and nonvascular structures within the kidney (1, 15), and thus different groups of renal nerves may exist that are
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influenced in an opposite manner by thoracic afferent nerves. Alternatively, the opposite responses may be consequent to simultaneous activation of afferent fibers having similar conduction velocities but different functions and different central synaptic connections. Sympathetic afferent neurons in the thorax originate from receptors in the heart, lungs, and great vessels (11,14,16, 22). Reflexes initiated from these areas under differing physiological conditions would not necessarily be ident cal. Thus the mixed renal nerve response to thoracic sympathetic afferent stimulation may be attributed to activation of a heterogeneous afferent population of neurons. A final interpretation of the biphasic character of this sympathosympathetic reflex is that renal nerve excitation and the following inhibition may not be separable responses and instead reflect an integrated response pattern of the central nervous system. The purpose of the present study was to investigate the possibility that the biphasic nature of renal nerve responses to electrical activation of thoracic afferent nerves can be attributed to heterogeneity within afferent or efferent components of the sympathetic reflex arc. METHODS
General Procedures Forty cats were anesthetized with intravenously administered a-chloralose (60 mg/kg). During the surgical preparation, cats were immobilized with gallamine triethiodide (4 mg/kg) to ensure muscle relaxation and were artificially respired. During an experiment, after the animal’s plane of anesthesia was assessed, supplemental doses of gallamine (l-2 mg/kg) were administered if needed. Blood gasesand pH were monitored and in some experiments animals were respired with 50% oxygen to maintain these variables within normal limits. Esophageal temperature was maintained at approximately 37OC. In all cats a femoral artery and vein were cannulated and a tracheostomy was performed. Carotid sinus and aortic arch baroreceptors and vagally innervated cardiopulmonary receptors were denervated by severing the vagus and glossopharyngeal nerves as they passed into the jugular foramen. The left kidney was exposed retroperitoneally and a small postganglionic renal nerve filament was prepared for recording electrical activity. Multifiber renal nerve activity was recorded with bipolar platinum electrodes using the standard electrophysiolog-
0363-6135/79/0000-0000$01.25
Copyright
0 1979 the American
Physiological
Society
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ical techniques described previously (25). During an experiment, neural discharge was monitored on an oscilloscope and recorded on magnetic tape for later analysis of spike frequencies with the use of a window discriminator and a Nicolet computer (Nicolet Instrument Corporation, Madison, WI). Mean neural discharge rate often was derived from a rate analyzer (Frederick Haer & Co., Brunswick, ME) and displayed on a Grass polygraph during an experiment. Femoral arterial blood pressure was displayed on a Grass polygraph and also recorded on magnetic tape. Specific Procedures Single unit recordings of renal sympathetic responses to electrical stimulation of thoracic sympathetic afferent nerues. The left stellate ganglion was approached retropleurally in four cats by removing portions of the first and second ribs. A nerve bundle emanating from the caudal edge of the ganglion was tied and severed, and the central segment was prepared for electrical stimulation of afferent fibers. The nerve bundle was placed on a bipolar platinum electrode and stimulated with 0.5ms square-wave pulses from a Grass S48 stimulator. Stimuli were delivered as lo-ms trains of three equally spaced pulses that varied in intensity between 2 and 12 V. One IO-ms train of three pulses was delivered every 1 or 2 s. Responses of individual postganglionic renal efferent fibers to this afferent stimulation were examined. Renal nerves were desheathed and split on a small dental mirror until activity of only a few fibers could be recorded. Movement artifacts in neural recordings were minimized by a pneumothoracotomy. Unit activity was recorded on magnetic tape and, with the use of a window discriminator, poststimulus histograms of individual unit responses were constructed by a Nicolet computer and plotted with an X-Y recorder. Multifiber renal sympathetic responses to activation of cardiac stretch receptors. Responses of multifiber renal nerves to stretch of specific regions of the heart were ascertained in nine cats by a technique devised to selectively stretch limited areas of the myocardium without dramatically changing systemic, pulmonary, or intracardiac pressures. Two rows of three individual sutures were placed in the ventricles, and several sutures were placed in the atria. The rows of ventricular sutures were 3 cm apart and the atria1 sutures also were placed 3 cm apart. Manual traction on these sutures was used to stretch the myocardium between them, thus activating stretch receptors in the area. Approximately 45 g/cm2 of force (measured with a Grass force-displacement transducer) were applied to the left ventricle, 30 g/cm2 to the right ventricle, and 40 g/cm2 to the atria. Multifiber renal nerve responses in these and the following experiments were monitored and analyzed as described in General Procedures. In addition to systemic blood pressure, atrial and/or ventricular pressures were monitored in seven cats. In most cats responses to cardiac stretch were compared before and after painting the surface of the stretched area with 10% phenol. Multifiber renal nerve responses to activation of stretch receptors in the thoracic aorta. Renal nerve responses to stretch of the aorta were observed in 11 cats
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after placement of a balloon-cuffed metal cannula in the upper thoracic aorta. The cannula was 5.5 cm long with a 4.0-cm balloon cuff. Venous inflow to the heart was occluded temporarily and the distal thoracic aorta transected to allow placement of the cannula. The proximal tip of the cannula was positioned close to the arch of the aorta. Inflation of the balloon with saline stretched the aortic segment without obstructing flow to the lower thoracic aorta. Renal nerve responses to aortic stretch were observed before and after denervation of the cannulated region by stripping the aorta of nerves and connective tissue and painting it with 10% phenol. Multifiber renal nerve responses to stretch of pulmonary vascular receptors. An intravenous infusion of 3% dextran in saline was employed to expand vascular volume, thereby increasing the discharge rate of pulmonary and other vascular stretch receptors. A volume of 15 ml/ kg was infused at a rate of 4.4 ml/min. The pulmonary vessels were denervated in six cats by removal of surface connective tissue and topical application of 10% phenol. Each of the seven pulmonary veins and the pulmonary arteries distal to the bifurcation of the main artery were denervated. The atria and main pulmonary artery usually were not denervated. The main pulmonary artery was left intact to prevent damage to ventricular fibers that pass between the root of the aorta and pulmonary artery. Renal nerve responses to intravascular volume expansion in these cats were compared to responses of six cats whose pulmonary sympathetic nerves remained intact. Central venous pressure was monitored in all cats, and pulmonary venous pressure was recorded in six cats. In four more large cats, the pulmonary circulation was perfused separately to allow pulmonary vascular pressure to be increased selectively without accompanying changes in systemic and intracardiac pressure. The weight of these cats ranged between 4 and 7 kg. A pistontype pulsatile/blood pump (Harvard Apparatus Co., Millis, MA) was used and the perfusion circuit was primed with approximately 30 ml of 3% dextran and 30 ml of feline blood. Inflow to the pump was provided via a cannula positioned in the right atrium, and outflow from the pump was directed to a cannula placed initially in the right ventricle and later advanced into the pulmonary artery. These cannulas were constructed of flexible plastic tubing with an outside diameter of 6 mm. Pulmonary venous outflow was partially occluded by a small balloon placed in the left atrium. Pressures were monitored in the right ventricle, main pulmonary artery, pulmonary vein, and femoral artery. The lungs were perfused at rates varying between 250 and 500 ml/min. The goal of the experiment was to increase pulmonary vascular pressures while maintaining systemic pressure constant and right ventricular pressure equal to or less than the control pressure. In several cats, increases in right ventricular pressure were prevented by placing a cannula in the ventricle that drained into the pump reservoir. Renal nerve activity was compared in each animal during low and high pulmonary pressures. Statistical Analysis of Data Changes in renal nerve activity during cardiac stretch were verified with an analysis of variance. Mean values
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were compared with a Student-Newman-Keuls test (20). Variability within a group was expressed as a coefficient of variability. Differences were considered statistically significant when P < 0.05. Changes in renal nerve activity and venous pressure during intravascular volume expansion were considered significant if they exceeded the limits of a 95% confidence interval (Z t S;) (20).
renal nerve activity, whereas increasing pulmonary vascular pressure inhibited activity. However, these techniques altered pressure nonselectively in the heart and thoracic circulation and caused changes in systemic blood pressure; therefore, these experiments could not be considered conclusive, and a selective technique (described in METHODS) was employed to activate different groups of cardiac stretch receptors. Stretch of both right and left ventricles increased renal RESULTS nerve activity. As shown in Fig. 2, renal nerve activity The responses of single postganglionic renal efferent was increased approximately 50% by stretch of the right neurons to electrical stimulation of thoracic sympathetic ventricle, while systemic blood pressure was decreased afferent nerves were examined to determine individual slightly. When blood pressure was decreased similarly by response patterns. As shown in Fig. 1, renal neurons hemorrhage in these cats, equivalent changes in renal responded to afferent stimulation with excitation fol- nerve activity did not occur. Reflex renal nerve responses lowed by inhibition of spontaneous discharge. The his- to stretch of the left ventricle usually were not accomtograms of activity show that this pattern occurred con- panied by changes in systemic blood pressure. During 30 sistently. Excitation occurred with a latency of approxior 60 s of right or left ventricular stretch, nerve activity mately 90 ms and the period of inhibition lasted from 150 increased in each of seven cats, and after release of to 1,500 ms. The excitation and following inhibition could stretch, activity promptly returned to control (Fig. 4). not be dissociated by altering stimulus intensity. The Mean activity during stretch was significantly greater silent period observed in some neurons was brief, indithan control activity. cating that inhibitory influences on these cells were minIncreases in renal nerve activity were shown to be reflexly mediated via sympathetic afferent nerves in five imal. No evidence could be ,obtained suggesting that excitation occurred predominantly in neurons that were cats. After initial documentation of the excitatory renormally quiescent whereas inhibitory responses oc- sponse, the area to be stretched was denervated with curred in spontaneously active neurons. Usually, when a topically applied phenol. The response, which was illusneuron responded to afferent stimulation, it displayed trated in Fig. 2, was abolished by this denervation (Fig. the biphasic pattern; however, an occasional unit ap- 3). In some cats, the response was not completely elimipeared to be only excited or only inhibited and some nated by this procedure. However, since phenol does not renal units did not respond to stimulation. penetrate through the entire myocardium, denervation Because most individual renal neurons were sequenprobably was not complete in many of the animals. tially excited and then inhibited by electrical stimulation In contrast, stretch of the atria produced no change in of the cardiopulmonary afferent nerves, the possibility renal nerve activity, as illustrated in Fig. 4. This may be was considered that different groups of afferent fibers interpreted as a failure of the technique to adequately were responsible for each of the two responses. These excite the receptors or as evidence that sympathetic potential groups could not be discriminated easily by afferent reflexes from the atria have little influence on differences in threshold to electrical stimulation. Thererenal nerve activity. fore, more natural stimuli were used in an attempt to Because stretch of the aorta excites sympathetic afferselectively activate afferent fibers responsible for excitent fibers and can initiate reflex changes in thoracic atory or inhibitory responses. An initial hypothesis was sympathetic efferent activity (18,22), the effect of stretch that these different fibers might originate from different of this region on renal nerve activity was determined. anatomic regions of the thoracic viscera. The renal nerve responses to aortic stretch in seven cats In preliminary experiments, ligatures were used to are shown in Fig. 5. In six of these cats, activation of occlude the great vessels, or balloons were distended aortic sympathetic afferent fibers caused reflex excitation within the heart or great vessels. In these experiments, it of renal nerve activity, which did not occur after the appeared that increasing intracardiac pressure increased stretched aortic segment had be.en denervated. During t
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FIG. 1. Single unit renal nerve responses to electrical stimulation of afferent fibers in the inferior cardiac nerve. A: spontaneous and poststimulus activity of a renal efferent fiber. B: histograms constructed from 50 samples of spontaneous or poststimulus activity. All poststimulus records were triggered by the application of 10 V, 0.5ms stimuli to the severed afferent nerve. The stimuli were delivered once every 2 s as lo-ms trains of 3 equally spaced pulses. The time of application of this stimulus is denoted on the records by a dot. An individual renal fiber that was active spontaneously responded predictably to this stimulation with excitation followed by inhibition.
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expansion is caused by activation of sympathetic afferent fibers from pulmonary vessels. To test this hypothesis, responses to expansion of intravascular volume were compared in animals with intact vs. denervated pulmonary vessels. Central venous pressure was monitored in all cats, and pulmonary venous pressure was monitored as well in six cats. Pulmonary venous pressure increased in parallel with central venous pressure during intravascular volume expansion. Changes in these pressures were very similar in magnitude and time course. As the intravenous infusion caused central venous pressure to increase, renal nerve activity was inhibited in cats with intact pulmonary sympathetic nerves (Fig. 6). In contrast, nerve activity did not change significantly in cats with denervated pulmonary blood vessels, although venous pressure increased similarly. Thus, inhibition of renal nerve activity during volume expansion appeared to be caused by activation of sympathetic afferent fibers from pulmonary vessels. In another group of experiments designed to test the same hypothesis, pulmonary vascular pressure was increased selectively with a perfusion pump without causing major changes in other cardiac or systemic pressures.
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180 3 2. Renal nerve and systemic blood pressure response to stretch of the right ventricle. A: mean discharge rate of a renal nerve derived from a rate analyzer and displayed on a polygraph simultaneously with arterial blood pressure. Time in seconds is indicated under the blood pressure trace. Times of initiation and release of stretch are marked on the time base. B: representative records of renal nerve activity during control (I), stretch (Z), and recovery (3) periods. Vertical calibration is 100 pV. Numbers above mean discharge tracing in A indicate approximate time interval during which these representative samples were taken. Stretch of the right ventricle increased renal nerve activity markedly and caused a small decrease in systemic blood pressure. Upon release of stretch, renal nerve activity and blood pressure returned promptly to control. FIG.
aortic stretch, blood pressure increased by 10-65 mmHg, a response that also was blocked by denervation. Mean carotid and . femoral arterial blood pressures in these cats were 93 and 81 mmHg, respectively. Four cats were not responsive to aortic stretch and are not includ .ed in the data reported in Fig. 5. The procedures involved in placement of the cannula caused deterioration of the animal’s condition in some experiments, that may have been a critical factor in the failure of these animals to demonstrate this reflex. Because it was not p ossible to group and quantify responses from all of the cats, the results were not analyzed statistically. The influences of pulmonary vascular sympathetic afferent fibers on renal nerves were ascertained next. These afferent neurons are excited by intravascular volume expansion (11, 16), which also causes inhibition of renal nerve activity (25). Therefore, the possibility was considered that inhibition of renal nerve activity during volume
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Due to the technical difficulties of these experiments, this criterion could be met only in four of many experiments. The responses of these four cats are shown in Fig. 7. When pulmonary arterial and venous pressures were increased, renal nerve activity decreased. These data also support the suggestion that inhibitory influences of thoracic sympathetic afferent neurons on renal efferent ac-
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7. Renal nerve response to selectively increased pulmonary venous (MPV) and arterial (MPA) pressure. Each line represents average renal nerve activity or mean blood pressure during a 30 to 60 s control (C) or experimental (E) sampling period in one cat. Responses of each of the four cats are denoted with a different symbol. As pulmonary pressures were increased, renal nerve activity was inhibited while mean femoral arterial (MFA) and right ventricular systolic (RV,& pressures remained relatively constant. FIG.
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DISCUSSION
Few definitive data are available clarifying the possible reflex influences of cardiopulmonary sympathetic afferent nerves on the kidney. Electrical stimulation of thoracic sympathetic afferent nerves has been shown to produce an excitation of renal nerve activity followed by inhibition of discharge (25). This experimental observation documents potential influences of these afferent fibers on sympathetic outflow to the kidney and suggests that physiological activation of these afferents may cause net excitation or inhibition of renal nerve activity. However, the neural organization of this complex response pattern and the circumstances under which net excitation or inhibition would be elicited are not known. Several hypotheses concerning the neural organization are consistent with this reflex pattern. When observed in multifiber neural recordings, excitation with following inhibition can reflect heterogeneity within the sampled neural efferent population. That is, some renal neurons may be excited by thoracic afferent nerves while the spontaneous discharge of others is inhibited. This type of response pattern occurs within other sympathetic reflex arcs. Noxious or nonnoxious stimulation of the skin elicits opposite responses in postganglionic fibers innervating cutaneous blood vessels and those innervating vasculature of muscles (4,5). Cardiac vagal afferent fibers have opposite influences on cardiac and renal efferent nerve activity (6). Since renal nerves innervate different renal structures (1, 15), opposing responses within two different populations of renal neurons would have been credible. However, in the present study individual renal neurons responded to afferent stimulation with both excitation and inhibition, illustrating that the biphasic responses observed in multiunit recordings could not be attributed solely to opposite responses of two groups within the renal efferent population. Because an exhaustive study of many renal units was not conducted, the possibility that some fibers can be excited while others are inhibited cannot be denied. In fact, a few fibers appeared to respond in this manner. An alternative hypothesis is that heterogeneity exists within the thoracic sympathetic afferent population of neurons: some afferents cause excitation of renal nerve activity while others produce inhibition of discharge. Excitatory reflex influences of thoracic sympathetic afferent fibers on blood pressure and heart rate are well known (10, 13, 19) and inhibitory influences” also have been described (12, 18). Pagani et al. (18) have shown that afferent fibers from the heart and aorta have converging and opposite influences on single upper thoracic preganglionic neurons. Thus, different sympathetic afferent fibers also might have opposite influences on renal efferent neurons. This possibility was explored extensively and confirmed in the present study. Stretch of the ventricles excited renal nerve activity via sympathetic afferent pathways. In contrast to the results of Pagani et al. (18), stretch of the aorta either had no effect on renal nerve activity or also caused an increased discharge.
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Thus, patterns of convergent sympathetic afferent influences onto lower thoracic preganglionic neurons apparently differ from those influences on upper thoracic neurons (10). In contrast, atria1 stretch failed to alter renal nerve activity. This lack of effect was predictable since other investigators have shown that atria1 reflex influences on renal nerves are mediated primarily (8) or solely (6) by the afferent vagus. Inhibitory sympathetic afferent influences on renal nerves were localized with much greater difficulty. A previous report had shown that intravascular volume expansion causes inhibition of renal nerve activity, a reflex effect that is dependent upon intact thoracic sympathetic afferent nerves (25). However, intravascular volume expansion increases activity in cardiac and aortic as well as pulmonary vascular sympathetic afferent fibers because pressure increases in high and low pressure areas of the circulation. Localization of receptors responsible for sympathoinhibition caused by volume expansion is possible only if reflex influences from one region predominate over influences from other regions. The failure of volume expansion to inhibit renal nerve activity after pulmonary denervation suggests that pulmonary sympathetic afferent fibers were responsible for the inhibition. Nishi et al. (16) and Lombardi et al. (11) have shown that sympathetic afferent fibers from the pulmonary arteries and veins discharge spontaneously at resting pressures and increase their firing steadily when intravascular volume is increased. These afferents may be those responsible for inhibition of renal nerve activity. This conclusion is subject to the criticism that the denervation technique used in the present study may have damaged fibers emerging from the ventricles; however, ventricular sympathetic afferent fibers were shown in the present study to have excitatory rather than inhibitory influences on renal nerve activity and would not have contributed to volume expansion-induced sympathoinhibition. The experiments in which pulmonary pressure was increased more selectively also lend credence to this hypothesis. When pulmonary vascular pressure was increased, renal nerve activity decreased. This reflex was seen in four cats that differed markedly with respect to initial intracardiac, intrapulmonary, and systemic arterial blood pressures. These results appear to differ from those of Ledsome and Kan (9), who increased pressure in isolated pulmonary artery pouches in dogs with intact pulmonary innervation and were unable to elicit changes in renal vascular resistance. However, inhibition of renal nerve activity, which may affect renal function significantly (2), does not necessarily alter vascular resistance. Thus, the results of Ledsome and Kan are not in conflict with those reported here. The present experiments do not define the magnitude of excitatory or inhibitory reflex influences that thoracic sympathetic afferent nerves may have on renal nerve activity. The sympathoinhibition evoked by these afferent nerves does not appear to be as dramatic as that which can be produced by carotid sinus (7) or left atria1 vagal (6) pressoreceptors. The excitation elicited from the ventricles or aorta also was not intense; however, receptors were activated only in a portion of these areas.
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H5o In contrast, true physiological activation probably would include more cardiac and aortic receptors and perhaps would evoke a greater response. An important consideration is that the sympathosympathetic reflexes act in concert with many other a&rent influences on central sympathetic neurons. Two influences that are difficult to reconcile initially are those of vagal and sympathetic afferent fibers originating from ventricular stretch receptors. The sympathetic afferent neurons cause reflex excitation of s$mpathetic activity, whereas ventricular vagal afferent fibers have been rkported to induce reflex hypckension (3), renal vasodilation (17), and decreased renin secretion (21), effects probably produced by inhibition of sympathetic tone. The most likely interpretation of this apparent paradox -~ is that these afferent neurons differ in receptor sensitivity or specific anatomic location or that the natural stimulus for either of the two groups may include factors other than stretch. When fully innervated ventricles are distended, the net effect appears to be inhibition of sympathetic tone (3). This suggests that ventricular vagal inhibitory influences are dominant over sympathetic excitatory influences when both groups mi activated by stretch. The most dramatic contribution of the sympathetic afferent neurons to contrul of the circulation may
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occur when they are sensitized ur activated by stimuli other than stretch. Several investigators have shown that chemical substances such as potassium (24) and bradykinin (23) can increase the discharge rate of cardiac sympathetic afferent fibers. The concentration of such substances in the myocardium could be altered in various circumstances. When bradykinin is applied tu the surface of the heart, profound pressor responses and excitation of renal nerve activity occur (26). These responses are caused by activation of cardiac sympathetic afferent nerves and can be observed when cardiac vagal innervation is intact (27). Thus, the present study has clarified the potential influences of these afferent nerves on sympathetic outflow to the kidney. Further investigation is needed to define the circumstances in which they have dominant influence on the kidney or &her components of central sympathetic outflow. The authors are indebted to Ms. Nancy Turner for expert assistance in the preparation of this manuscript. Special appreciation is extended to Drs, Peter Schwartz and Massimo Pagani for their instruction regarding the use of the aortic cannuta. This research was supported by a grant from the Michigan Heart Association and by Grant HL-21436 from the National He-art, Lung, and Blood Institute. Received 4 December 1978; accepted in final form 8 March 1979,
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G. F. Neural control of renal tubular sodium reabsorption in the dog. Federation Proc. 37: 1214~1217,1978. 3. Fox, I. J., D, A. GERASCH, AND J. J. LEONARD. Left ventricular mechanoreceptors: a hemodynamic study. J. physi&. London 273:
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G. RECURDATI, AND P. SCHWARTZ. Spinal sympathetic reflexes elicited by increases in arterial blood pressure. Am. J. Physiol. 220: 128-134, 1971, 13. MALLIANI, A., M. PARKS, R P, TUCKETT, AND A. M. BROWN. Reflex increases in heart rate elicited by stimulation of afferent cardiac sympathetic nerve fibers in the cat. Circ, Res, 32: 9-14,
1973. 14. MALLIANI,
A., G. RECORDATI, AND P, G. SCHWARTZ. Nervous activity of afferent cardiac sympathetic f&es with atriaf and ven-
tricular endings. J; Fhysbl. London 229: 457-469; 1973. J., AND L. BARAJAS. Electron microscopic and histochemical evidence for a tubular innervation in the renal cortex of the monkey. J. Wtrastruct, Res. 41: 533-549, 1972. NISHI, K,, M. SAKANASHI, AND F. TAKENAKA. Afferent fibers from pulmonary arterial baroreceptors in the left cardiac sympathetic nerve of the cat. J, physiol. London 240: 53-66, 1974, &ERG, B., AND P. THOR&N, Circulatory responses to stimulation of medullated and non-medullated afferents in the cardiac nerve in the cat. Acta Physiol. Stand. 87: 121-132, 1973. PAGANI, M., P. SCHWARTZ, R. BANKS, F. LOMBARDI, AND A. MALLIANI. Reflex responses of sympathetic preganghonic neurones initiated by different cardiovascular receptors in spinal animals. Brain Res. 68: 215.225,1974, PETERSON, D,, AND A. BROWN. Pressor reflexes produced by stimulation of afferent fibers in the cardiac sympathetic nerves of the cat. Circ. Res. 28: 605-610, 1971. SOKOL, R. R., AND F, J. ROHLF, Biometry. The Principles and Practice of Statistics in Biological Research. San Francisco: Freeman, 1969. THAMES, M. Reflex suppression of renin release by ventricular receptors with vagal afferents. Am. J. PhysioZ. 233: H181-H184, 1977 or Am. J, PhysioL: Heart Circ. Physiol. 2: HI819Hl84, 1977, UCHIDA, Y. Afferent aortic nerve fibers with their pathways in cardiac sympathetic nerves. Am. J. Physiol. 228: 990.995,1975. UCHIDA, Y., AND S, MURAO. Bradykinin-induced excitation of afferent cardiac sympathetic nerve fibers. Jpn. Heart J, 15: 84-91, 1974, UCHIDA, Y., AND 23. MURAO. Potassium-induced excitation of afferm ent cardiac sympathetic nerve fibers. Am. JI Physiol. 226: 603-607,
15. MILLER,
18.
19.
20.
21, 22. 23.
24.
1974.
25, WEAVER, L. C. Cardiopulmonary sympathetic afferent influences on rend nerve activity. Am. J. Physl’ol. 233: H592-H599, 1977 or Am. J. Physiol.: Heart Circ. PhysbZ. 2: H592-)3[599, 1977, 26. WEAVER, L. C. Renal nerve responses to activation of cardiac sympathetic nerves by coronary occlusion and by topical application of autocoids to the myocardium. Neurosci. Abstr, 4: 26, 1978. 27. WEAVERS L. C., AND K. A. REIMANN. Comparison of sympathetic and vagal cardiac afferent influences on renal nerves, Federation Proc. 38: 1201,1979,
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afferent
LYNNE C. WEAVER, LINDA J. MACKLEM, KEITH A. REIMANN, ROBERT L. MECKLER, AND ROBERT S. OEHL Department of Physiology, Michigan State University, East Lansing, Michigan
LYNNE C., LINDA J. MACKLEM, KEITH A. REIROBERT L. MECKLER, AND ROBERT S. OEHL. Organization of thoracic sympathetic afferent influences on renal nerve activity. Am. J. Physiol. 237(l): H44-H50, 1979 or Am. J. Physiol.: Heart Circ. Physiol. 6(l): H44-H50, 1979.-The specific influences of thoracic sympathetic afferent nerves on sympathetic outflow to the kidney have not been defined clearly, although preliminary experiments have shown that electrical stimulation of these afferent nerves causes excitation of multifiber renal nerve activity followed by inhibition of discharge. The organization of this complex reflex was investigated further in anesthetized, vagotomized, sinoaortic-denervated cats. The apparent ability of the cardiopulmonary afferent nerves to excite and then inhibit multifiber renal nerve activity initially was attributed to opposite responses of individual renal neurons to afferent stimulation. This hypothesis was not confirmed in the present study because many single postganglionic renal fibers responded to electrical afferent stimulation with excitation followed by inhibition of discharge. Alternatively, the possibility was considered that different thoracic sympathetic afferent neurons have opposite influences on renal nerve activity. In support of this suggestion, sympathetic afferent fibers activated by stretch of the ventricles or aorta caused reflex excitation of renal nerve activity whereas activation of afferent fibers from the pulmonary vasculature caused an inhibition of this discharge. Thus, the complexity of thoracic sympathetic afferent influences on sympathetic outflow to the kidney is likely related to heterogeneity within the afferent rather than efferent neural population. WEAVER,
MANN,
electrophysiological
analysis; reflex organization;
cat
nervescanhavecomplex influences on sympathetic outflow to the kidney. Activation of these afferent fibers by a brief electrical stimulus has been shown to evoke an excitatory burst in renal nerve activity followed by inhibition of discharge (25). The neural organization responsible for the biphasic character of this response is not understood. This response can be produced by activation of a homogeneous group of myelinated A6 fibers. Because the response was observed in multifiber recording of renal nerve activity, it may reflect heterogeneity within the renal efferent population. That is, activation of a group of similar sympathetic afferent fibers may excite some renal efferent neurons while inhibiting spontaneous activity in others. Sympathetic nerves innervate several vascular and nonvascular structures within the kidney (1, 15), and thus different groups of renal nerves may exist that are
THORACICSYMPATHETICAFFERENT
H44
48824
influenced in an opposite manner by thoracic afferent nerves. Alternatively, the opposite responses may be consequent to simultaneous activation of afferent fibers having similar conduction velocities but different functions and different central synaptic connections. Sympathetic afferent neurons in the thorax originate from receptors in the heart, lungs, and great vessels (11,14,16, 22). Reflexes initiated from these areas under differing physiological conditions would not necessarily be ident cal. Thus the mixed renal nerve response to thoracic sympathetic afferent stimulation may be attributed to activation of a heterogeneous afferent population of neurons. A final interpretation of the biphasic character of this sympathosympathetic reflex is that renal nerve excitation and the following inhibition may not be separable responses and instead reflect an integrated response pattern of the central nervous system. The purpose of the present study was to investigate the possibility that the biphasic nature of renal nerve responses to electrical activation of thoracic afferent nerves can be attributed to heterogeneity within afferent or efferent components of the sympathetic reflex arc. METHODS
General Procedures Forty cats were anesthetized with intravenously administered a-chloralose (60 mg/kg). During the surgical preparation, cats were immobilized with gallamine triethiodide (4 mg/kg) to ensure muscle relaxation and were artificially respired. During an experiment, after the animal’s plane of anesthesia was assessed, supplemental doses of gallamine (l-2 mg/kg) were administered if needed. Blood gasesand pH were monitored and in some experiments animals were respired with 50% oxygen to maintain these variables within normal limits. Esophageal temperature was maintained at approximately 37OC. In all cats a femoral artery and vein were cannulated and a tracheostomy was performed. Carotid sinus and aortic arch baroreceptors and vagally innervated cardiopulmonary receptors were denervated by severing the vagus and glossopharyngeal nerves as they passed into the jugular foramen. The left kidney was exposed retroperitoneally and a small postganglionic renal nerve filament was prepared for recording electrical activity. Multifiber renal nerve activity was recorded with bipolar platinum electrodes using the standard electrophysiolog-
0363-6135/79/0000-0000$01.25
Copyright
0 1979 the American
Physiological
Society
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THORACIC
SYMPATHETIC
AFFERENT
INFLUENCE
ON
RENAL
ical techniques described previously (25). During an experiment, neural discharge was monitored on an oscilloscope and recorded on magnetic tape for later analysis of spike frequencies with the use of a window discriminator and a Nicolet computer (Nicolet Instrument Corporation, Madison, WI). Mean neural discharge rate often was derived from a rate analyzer (Frederick Haer & Co., Brunswick, ME) and displayed on a Grass polygraph during an experiment. Femoral arterial blood pressure was displayed on a Grass polygraph and also recorded on magnetic tape. Specific Procedures Single unit recordings of renal sympathetic responses to electrical stimulation of thoracic sympathetic afferent nerues. The left stellate ganglion was approached retropleurally in four cats by removing portions of the first and second ribs. A nerve bundle emanating from the caudal edge of the ganglion was tied and severed, and the central segment was prepared for electrical stimulation of afferent fibers. The nerve bundle was placed on a bipolar platinum electrode and stimulated with 0.5ms square-wave pulses from a Grass S48 stimulator. Stimuli were delivered as lo-ms trains of three equally spaced pulses that varied in intensity between 2 and 12 V. One IO-ms train of three pulses was delivered every 1 or 2 s. Responses of individual postganglionic renal efferent fibers to this afferent stimulation were examined. Renal nerves were desheathed and split on a small dental mirror until activity of only a few fibers could be recorded. Movement artifacts in neural recordings were minimized by a pneumothoracotomy. Unit activity was recorded on magnetic tape and, with the use of a window discriminator, poststimulus histograms of individual unit responses were constructed by a Nicolet computer and plotted with an X-Y recorder. Multifiber renal sympathetic responses to activation of cardiac stretch receptors. Responses of multifiber renal nerves to stretch of specific regions of the heart were ascertained in nine cats by a technique devised to selectively stretch limited areas of the myocardium without dramatically changing systemic, pulmonary, or intracardiac pressures. Two rows of three individual sutures were placed in the ventricles, and several sutures were placed in the atria. The rows of ventricular sutures were 3 cm apart and the atria1 sutures also were placed 3 cm apart. Manual traction on these sutures was used to stretch the myocardium between them, thus activating stretch receptors in the area. Approximately 45 g/cm2 of force (measured with a Grass force-displacement transducer) were applied to the left ventricle, 30 g/cm2 to the right ventricle, and 40 g/cm2 to the atria. Multifiber renal nerve responses in these and the following experiments were monitored and analyzed as described in General Procedures. In addition to systemic blood pressure, atrial and/or ventricular pressures were monitored in seven cats. In most cats responses to cardiac stretch were compared before and after painting the surface of the stretched area with 10% phenol. Multifiber renal nerve responses to activation of stretch receptors in the thoracic aorta. Renal nerve responses to stretch of the aorta were observed in 11 cats
NERVES
H45
after placement of a balloon-cuffed metal cannula in the upper thoracic aorta. The cannula was 5.5 cm long with a 4.0-cm balloon cuff. Venous inflow to the heart was occluded temporarily and the distal thoracic aorta transected to allow placement of the cannula. The proximal tip of the cannula was positioned close to the arch of the aorta. Inflation of the balloon with saline stretched the aortic segment without obstructing flow to the lower thoracic aorta. Renal nerve responses to aortic stretch were observed before and after denervation of the cannulated region by stripping the aorta of nerves and connective tissue and painting it with 10% phenol. Multifiber renal nerve responses to stretch of pulmonary vascular receptors. An intravenous infusion of 3% dextran in saline was employed to expand vascular volume, thereby increasing the discharge rate of pulmonary and other vascular stretch receptors. A volume of 15 ml/ kg was infused at a rate of 4.4 ml/min. The pulmonary vessels were denervated in six cats by removal of surface connective tissue and topical application of 10% phenol. Each of the seven pulmonary veins and the pulmonary arteries distal to the bifurcation of the main artery were denervated. The atria and main pulmonary artery usually were not denervated. The main pulmonary artery was left intact to prevent damage to ventricular fibers that pass between the root of the aorta and pulmonary artery. Renal nerve responses to intravascular volume expansion in these cats were compared to responses of six cats whose pulmonary sympathetic nerves remained intact. Central venous pressure was monitored in all cats, and pulmonary venous pressure was recorded in six cats. In four more large cats, the pulmonary circulation was perfused separately to allow pulmonary vascular pressure to be increased selectively without accompanying changes in systemic and intracardiac pressure. The weight of these cats ranged between 4 and 7 kg. A pistontype pulsatile/blood pump (Harvard Apparatus Co., Millis, MA) was used and the perfusion circuit was primed with approximately 30 ml of 3% dextran and 30 ml of feline blood. Inflow to the pump was provided via a cannula positioned in the right atrium, and outflow from the pump was directed to a cannula placed initially in the right ventricle and later advanced into the pulmonary artery. These cannulas were constructed of flexible plastic tubing with an outside diameter of 6 mm. Pulmonary venous outflow was partially occluded by a small balloon placed in the left atrium. Pressures were monitored in the right ventricle, main pulmonary artery, pulmonary vein, and femoral artery. The lungs were perfused at rates varying between 250 and 500 ml/min. The goal of the experiment was to increase pulmonary vascular pressures while maintaining systemic pressure constant and right ventricular pressure equal to or less than the control pressure. In several cats, increases in right ventricular pressure were prevented by placing a cannula in the ventricle that drained into the pump reservoir. Renal nerve activity was compared in each animal during low and high pulmonary pressures. Statistical Analysis of Data Changes in renal nerve activity during cardiac stretch were verified with an analysis of variance. Mean values
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H46
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ET AL.
were compared with a Student-Newman-Keuls test (20). Variability within a group was expressed as a coefficient of variability. Differences were considered statistically significant when P < 0.05. Changes in renal nerve activity and venous pressure during intravascular volume expansion were considered significant if they exceeded the limits of a 95% confidence interval (Z t S;) (20).
renal nerve activity, whereas increasing pulmonary vascular pressure inhibited activity. However, these techniques altered pressure nonselectively in the heart and thoracic circulation and caused changes in systemic blood pressure; therefore, these experiments could not be considered conclusive, and a selective technique (described in METHODS) was employed to activate different groups of cardiac stretch receptors. Stretch of both right and left ventricles increased renal RESULTS nerve activity. As shown in Fig. 2, renal nerve activity The responses of single postganglionic renal efferent was increased approximately 50% by stretch of the right neurons to electrical stimulation of thoracic sympathetic ventricle, while systemic blood pressure was decreased afferent nerves were examined to determine individual slightly. When blood pressure was decreased similarly by response patterns. As shown in Fig. 1, renal neurons hemorrhage in these cats, equivalent changes in renal responded to afferent stimulation with excitation fol- nerve activity did not occur. Reflex renal nerve responses lowed by inhibition of spontaneous discharge. The his- to stretch of the left ventricle usually were not accomtograms of activity show that this pattern occurred con- panied by changes in systemic blood pressure. During 30 sistently. Excitation occurred with a latency of approxior 60 s of right or left ventricular stretch, nerve activity mately 90 ms and the period of inhibition lasted from 150 increased in each of seven cats, and after release of to 1,500 ms. The excitation and following inhibition could stretch, activity promptly returned to control (Fig. 4). not be dissociated by altering stimulus intensity. The Mean activity during stretch was significantly greater silent period observed in some neurons was brief, indithan control activity. cating that inhibitory influences on these cells were minIncreases in renal nerve activity were shown to be reflexly mediated via sympathetic afferent nerves in five imal. No evidence could be ,obtained suggesting that excitation occurred predominantly in neurons that were cats. After initial documentation of the excitatory renormally quiescent whereas inhibitory responses oc- sponse, the area to be stretched was denervated with curred in spontaneously active neurons. Usually, when a topically applied phenol. The response, which was illusneuron responded to afferent stimulation, it displayed trated in Fig. 2, was abolished by this denervation (Fig. the biphasic pattern; however, an occasional unit ap- 3). In some cats, the response was not completely elimipeared to be only excited or only inhibited and some nated by this procedure. However, since phenol does not renal units did not respond to stimulation. penetrate through the entire myocardium, denervation Because most individual renal neurons were sequenprobably was not complete in many of the animals. tially excited and then inhibited by electrical stimulation In contrast, stretch of the atria produced no change in of the cardiopulmonary afferent nerves, the possibility renal nerve activity, as illustrated in Fig. 4. This may be was considered that different groups of afferent fibers interpreted as a failure of the technique to adequately were responsible for each of the two responses. These excite the receptors or as evidence that sympathetic potential groups could not be discriminated easily by afferent reflexes from the atria have little influence on differences in threshold to electrical stimulation. Thererenal nerve activity. fore, more natural stimuli were used in an attempt to Because stretch of the aorta excites sympathetic afferselectively activate afferent fibers responsible for excitent fibers and can initiate reflex changes in thoracic atory or inhibitory responses. An initial hypothesis was sympathetic efferent activity (18,22), the effect of stretch that these different fibers might originate from different of this region on renal nerve activity was determined. anatomic regions of the thoracic viscera. The renal nerve responses to aortic stretch in seven cats In preliminary experiments, ligatures were used to are shown in Fig. 5. In six of these cats, activation of occlude the great vessels, or balloons were distended aortic sympathetic afferent fibers caused reflex excitation within the heart or great vessels. In these experiments, it of renal nerve activity, which did not occur after the appeared that increasing intracardiac pressure increased stretched aortic segment had be.en denervated. During t
l
916
SPONTANEOUS
POST-STIMULUS
-- -
FIG. 1. Single unit renal nerve responses to electrical stimulation of afferent fibers in the inferior cardiac nerve. A: spontaneous and poststimulus activity of a renal efferent fiber. B: histograms constructed from 50 samples of spontaneous or poststimulus activity. All poststimulus records were triggered by the application of 10 V, 0.5ms stimuli to the severed afferent nerve. The stimuli were delivered once every 2 s as lo-ms trains of 3 equally spaced pulses. The time of application of this stimulus is denoted on the records by a dot. An individual renal fiber that was active spontaneously responded predictably to this stimulation with excitation followed by inhibition.
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THORACIC
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AFFERENT
INFLUENCE
ON
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A 200 160 120
1
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H47
NERVES
expansion is caused by activation of sympathetic afferent fibers from pulmonary vessels. To test this hypothesis, responses to expansion of intravascular volume were compared in animals with intact vs. denervated pulmonary vessels. Central venous pressure was monitored in all cats, and pulmonary venous pressure was monitored as well in six cats. Pulmonary venous pressure increased in parallel with central venous pressure during intravascular volume expansion. Changes in these pressures were very similar in magnitude and time course. As the intravenous infusion caused central venous pressure to increase, renal nerve activity was inhibited in cats with intact pulmonary sympathetic nerves (Fig. 6). In contrast, nerve activity did not change significantly in cats with denervated pulmonary blood vessels, although venous pressure increased similarly. Thus, inhibition of renal nerve activity during volume expansion appeared to be caused by activation of sympathetic afferent fibers from pulmonary vessels. In another group of experiments designed to test the same hypothesis, pulmonary vascular pressure was increased selectively with a perfusion pump without causing major changes in other cardiac or systemic pressures.
A 2
NA
1
2
3
180 3 2. Renal nerve and systemic blood pressure response to stretch of the right ventricle. A: mean discharge rate of a renal nerve derived from a rate analyzer and displayed on a polygraph simultaneously with arterial blood pressure. Time in seconds is indicated under the blood pressure trace. Times of initiation and release of stretch are marked on the time base. B: representative records of renal nerve activity during control (I), stretch (Z), and recovery (3) periods. Vertical calibration is 100 pV. Numbers above mean discharge tracing in A indicate approximate time interval during which these representative samples were taken. Stretch of the right ventricle increased renal nerve activity markedly and caused a small decrease in systemic blood pressure. Upon release of stretch, renal nerve activity and blood pressure returned promptly to control. FIG.
aortic stretch, blood pressure increased by 10-65 mmHg, a response that also was blocked by denervation. Mean carotid and . femoral arterial blood pressures in these cats were 93 and 81 mmHg, respectively. Four cats were not responsive to aortic stretch and are not includ .ed in the data reported in Fig. 5. The procedures involved in placement of the cannula caused deterioration of the animal’s condition in some experiments, that may have been a critical factor in the failure of these animals to demonstrate this reflex. Because it was not p ossible to group and quantify responses from all of the cats, the results were not analyzed statistically. The influences of pulmonary vascular sympathetic afferent fibers on renal nerves were ascertained next. These afferent neurons are excited by intravascular volume expansion (11, 16), which also causes inhibition of renal nerve activity (25). Therefore, the possibility was considered that inhibition of renal nerve activity during volume
140 100 cts/bc
1
FIG. 3. Renal nerve and systemic blood pressure responses to stretch of right ventricle after denervation of stretched area of myocardium. Format is same as that of Fig. 2. Responses are from the cat illustrated in Fig. 2 after stretched area of right ventricle was painted with 10% phenol. Stretch of this ventricle no longer elicited any reflex change in renal nerve activity.
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H48
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FIG. 6. Changes in renal nerve activity and central venous pressure induced by intravascular volume expansion in cats with intact (closed symbols) or denervated (open symbols) pulmonary vasculature. Renal nerve activity is expressed as percent of control spontaneous discharge, and venous pressure is expressed as change from control. Data are shown as means -+ SE in each group of 6 cats. Infusion of 3% dextran in saline at 4.4 ml/min was begun at time zero and completed at 16 min. Total volume infused equaled 15 ml/kg. Asterisks indicate values not included within 95% confidence interval S;) around 100% renal nerve activity or zero change in venous pressure. Intravascular volume expansion increased venous pressure equivalently in both groups of cats and inhibited renal nerve activity only in cats with denervated pulmonary vasculature. (t
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Due to the technical difficulties of these experiments, this criterion could be met only in four of many experiments. The responses of these four cats are shown in Fig. 7. When pulmonary arterial and venous pressures were increased, renal nerve activity decreased. These data also support the suggestion that inhibitory influences of thoracic sympathetic afferent neurons on renal efferent ac-
m-ml)
C
E
C
E
7. Renal nerve response to selectively increased pulmonary venous (MPV) and arterial (MPA) pressure. Each line represents average renal nerve activity or mean blood pressure during a 30 to 60 s control (C) or experimental (E) sampling period in one cat. Responses of each of the four cats are denoted with a different symbol. As pulmonary pressures were increased, renal nerve activity was inhibited while mean femoral arterial (MFA) and right ventricular systolic (RV,& pressures remained relatively constant. FIG.
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THORACIC
SYMPATHETIC
tivity originate blood vessels.
AFFERENT
from stretch
receptors
INFLUENCE
ON
RENAL
in the pulmonary
DISCUSSION
Few definitive data are available clarifying the possible reflex influences of cardiopulmonary sympathetic afferent nerves on the kidney. Electrical stimulation of thoracic sympathetic afferent nerves has been shown to produce an excitation of renal nerve activity followed by inhibition of discharge (25). This experimental observation documents potential influences of these afferent fibers on sympathetic outflow to the kidney and suggests that physiological activation of these afferents may cause net excitation or inhibition of renal nerve activity. However, the neural organization of this complex response pattern and the circumstances under which net excitation or inhibition would be elicited are not known. Several hypotheses concerning the neural organization are consistent with this reflex pattern. When observed in multifiber neural recordings, excitation with following inhibition can reflect heterogeneity within the sampled neural efferent population. That is, some renal neurons may be excited by thoracic afferent nerves while the spontaneous discharge of others is inhibited. This type of response pattern occurs within other sympathetic reflex arcs. Noxious or nonnoxious stimulation of the skin elicits opposite responses in postganglionic fibers innervating cutaneous blood vessels and those innervating vasculature of muscles (4,5). Cardiac vagal afferent fibers have opposite influences on cardiac and renal efferent nerve activity (6). Since renal nerves innervate different renal structures (1, 15), opposing responses within two different populations of renal neurons would have been credible. However, in the present study individual renal neurons responded to afferent stimulation with both excitation and inhibition, illustrating that the biphasic responses observed in multiunit recordings could not be attributed solely to opposite responses of two groups within the renal efferent population. Because an exhaustive study of many renal units was not conducted, the possibility that some fibers can be excited while others are inhibited cannot be denied. In fact, a few fibers appeared to respond in this manner. An alternative hypothesis is that heterogeneity exists within the thoracic sympathetic afferent population of neurons: some afferents cause excitation of renal nerve activity while others produce inhibition of discharge. Excitatory reflex influences of thoracic sympathetic afferent fibers on blood pressure and heart rate are well known (10, 13, 19) and inhibitory influences” also have been described (12, 18). Pagani et al. (18) have shown that afferent fibers from the heart and aorta have converging and opposite influences on single upper thoracic preganglionic neurons. Thus, different sympathetic afferent fibers also might have opposite influences on renal efferent neurons. This possibility was explored extensively and confirmed in the present study. Stretch of the ventricles excited renal nerve activity via sympathetic afferent pathways. In contrast to the results of Pagani et al. (18), stretch of the aorta either had no effect on renal nerve activity or also caused an increased discharge.
NERVES
H49
Thus, patterns of convergent sympathetic afferent influences onto lower thoracic preganglionic neurons apparently differ from those influences on upper thoracic neurons (10). In contrast, atria1 stretch failed to alter renal nerve activity. This lack of effect was predictable since other investigators have shown that atria1 reflex influences on renal nerves are mediated primarily (8) or solely (6) by the afferent vagus. Inhibitory sympathetic afferent influences on renal nerves were localized with much greater difficulty. A previous report had shown that intravascular volume expansion causes inhibition of renal nerve activity, a reflex effect that is dependent upon intact thoracic sympathetic afferent nerves (25). However, intravascular volume expansion increases activity in cardiac and aortic as well as pulmonary vascular sympathetic afferent fibers because pressure increases in high and low pressure areas of the circulation. Localization of receptors responsible for sympathoinhibition caused by volume expansion is possible only if reflex influences from one region predominate over influences from other regions. The failure of volume expansion to inhibit renal nerve activity after pulmonary denervation suggests that pulmonary sympathetic afferent fibers were responsible for the inhibition. Nishi et al. (16) and Lombardi et al. (11) have shown that sympathetic afferent fibers from the pulmonary arteries and veins discharge spontaneously at resting pressures and increase their firing steadily when intravascular volume is increased. These afferents may be those responsible for inhibition of renal nerve activity. This conclusion is subject to the criticism that the denervation technique used in the present study may have damaged fibers emerging from the ventricles; however, ventricular sympathetic afferent fibers were shown in the present study to have excitatory rather than inhibitory influences on renal nerve activity and would not have contributed to volume expansion-induced sympathoinhibition. The experiments in which pulmonary pressure was increased more selectively also lend credence to this hypothesis. When pulmonary vascular pressure was increased, renal nerve activity decreased. This reflex was seen in four cats that differed markedly with respect to initial intracardiac, intrapulmonary, and systemic arterial blood pressures. These results appear to differ from those of Ledsome and Kan (9), who increased pressure in isolated pulmonary artery pouches in dogs with intact pulmonary innervation and were unable to elicit changes in renal vascular resistance. However, inhibition of renal nerve activity, which may affect renal function significantly (2), does not necessarily alter vascular resistance. Thus, the results of Ledsome and Kan are not in conflict with those reported here. The present experiments do not define the magnitude of excitatory or inhibitory reflex influences that thoracic sympathetic afferent nerves may have on renal nerve activity. The sympathoinhibition evoked by these afferent nerves does not appear to be as dramatic as that which can be produced by carotid sinus (7) or left atria1 vagal (6) pressoreceptors. The excitation elicited from the ventricles or aorta also was not intense; however, receptors were activated only in a portion of these areas.
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H5o In contrast, true physiological activation probably would include more cardiac and aortic receptors and perhaps would evoke a greater response. An important consideration is that the sympathosympathetic reflexes act in concert with many other a&rent influences on central sympathetic neurons. Two influences that are difficult to reconcile initially are those of vagal and sympathetic afferent fibers originating from ventricular stretch receptors. The sympathetic afferent neurons cause reflex excitation of s$mpathetic activity, whereas ventricular vagal afferent fibers have been rkported to induce reflex hypckension (3), renal vasodilation (17), and decreased renin secretion (21), effects probably produced by inhibition of sympathetic tone. The most likely interpretation of this apparent paradox -~ is that these afferent neurons differ in receptor sensitivity or specific anatomic location or that the natural stimulus for either of the two groups may include factors other than stretch. When fully innervated ventricles are distended, the net effect appears to be inhibition of sympathetic tone (3). This suggests that ventricular vagal inhibitory influences are dominant over sympathetic excitatory influences when both groups mi activated by stretch. The most dramatic contribution of the sympathetic afferent neurons to contrul of the circulation may
WEAVER
ET AL.
occur when they are sensitized ur activated by stimuli other than stretch. Several investigators have shown that chemical substances such as potassium (24) and bradykinin (23) can increase the discharge rate of cardiac sympathetic afferent fibers. The concentration of such substances in the myocardium could be altered in various circumstances. When bradykinin is applied tu the surface of the heart, profound pressor responses and excitation of renal nerve activity occur (26). These responses are caused by activation of cardiac sympathetic afferent nerves and can be observed when cardiac vagal innervation is intact (27). Thus, the present study has clarified the potential influences of these afferent nerves on sympathetic outflow to the kidney. Further investigation is needed to define the circumstances in which they have dominant influence on the kidney or &her components of central sympathetic outflow. The authors are indebted to Ms. Nancy Turner for expert assistance in the preparation of this manuscript. Special appreciation is extended to Drs, Peter Schwartz and Massimo Pagani for their instruction regarding the use of the aortic cannuta. This research was supported by a grant from the Michigan Heart Association and by Grant HL-21436 from the National He-art, Lung, and Blood Institute. Received 4 December 1978; accepted in final form 8 March 1979,
REFERENCES 1. BARAJAS, L., AND J. M%LER. The innervation of the juxtaglomerular apparatus and surrounding tubules: a quantitative analysis by serial section electron microscopy. J. Uttrastruct. Res. 43: 107-132, 1973. 2. DIBONA,
G. F. Neural control of renal tubular sodium reabsorption in the dog. Federation Proc. 37: 1214~1217,1978. 3. Fox, I. J., D, A. GERASCH, AND J. J. LEONARD. Left ventricular mechanoreceptors: a hemodynamic study. J. physi&. London 273:
16.
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