573
Journal of Physiology (1991), 432, pp. 573-584 With 2 figures Printed in Great Britain
EFFECT OF SOMATIC NERVE STIMULATION ON THE KIDNEY IN INTACT, VAGOTOMIZED AND CAROTID SINUS-DENERVATED RATS
BY GERARD DAVIS AND EDWARD J. JOHNS From the Department of Physiology, The Medical School, Birmingham B15 2TT
(Received 6 March 1990) SUMMARY
1. The influence of cardiopulmonary and arterial baroreceptors on the renal nervedependent functional responses of the kidney to electrical stimulation of somatic afferent nerves was studied in pentobarbitone-anaesthetized rats. 2. Electrical stimulation of the left brachial nerve plexus at 3 Hz, 0-2 ms and 15 V in the intact animals increased blood pressure by 22 %, and while renal perfusion pressure was maintained at pre-stimulus levels, renal blood flow and glomerular filtration rate decreased by 14 and 22 % respectively. At the same time urine flow rate and absolute and fractional sodium excretion decreased by 36, 42 and 27 % respectively. In animals subjected to acute renal nerve section these renal functional responses could not be elicited. 3. Following bilateral vagotomy the systemic and renal haemodynamic responses to brachial nerve stimulation were similar to the intact group. However, urine flow rate and absolute and fractional sodium excretions decreased by 50, 59 and 47 % respectively, responses which were significantly greater than in the intact group. 4. In a group of rats in which the carotid sinus nerves had been sectioned, stimulation of the brachial plexus caused reductions of renal blood flow and glomerular filtration rate of the same magnitude as in the intact group; however, urine flow rate and absolute and fractional sodium excretion fell by 51, 60 and 48%, respectively, which were significantly larger than in the intact group. 5. These results demonstrate that the afferent nerve information arising from muscle joints and skin and carried via the brachial plexus caused reflex renal nervedependent reductions in renal haemodynamics and an antidiuresis and antinatriuresis. The cardiopulmonary and carotid sinus baroreceptors exert a tonic inhibitory action on these reflex renal responses insofar as they appeared to attenuate the antidiuretic and antinatriuretic responses to somatic afferent nerve stimulation. INTRODUCTION
The exercise pressor reflex has both peripheral and central components (Mitchell, 1985). The peripheral limb of the reflex is initiated by activation of group IV, somatic afferent nerve fibres, arising from sensory receptors in skin, joints and muscle, which discharge into group III and IV 'somatic' afferent nerves (Coote, Hilton & PerezGonzalez, 1971; Mitchell, Kaufman & Iwamoto, 1983), resulting in an increased MS 8330
574
G. DAVIS ANDE. J. JOHNS
sympathetic nerve activity, which functionally increases heart rate (Sato & Schmidt, 1987) and peripheral resistance (Thames & Abboud, 1979). With regard to the kidney, Abboud, Mark & Thames (1981) showed that brief periods of high-frequency stimulation of the somatic afferent fibres contained within the sciatic nerve induced a profound decrease in renal blood flow which was approximately frequency related. Handa & Johns (1987), using the rat, found that stimulation of the somatic sensory fibres of the brachial nerve plexus at much lower frequencies increased renal sympathetic nerve activity which was associated with only minor falls in renal haemodynamics but large decreases in the excretion of sodium. At the level of the central nervous system a number of interactions may occur. It is now clear that the pressure receptors of the cardiovascular system can influence renal sympathetic nerve activity. Karim, Kidd, Malpus & Penna (1972) demonstrated that raising left atrial pressure in the dog, by inflating a small balloon in the pulmonary vein-left atrial junction, resulted in a decreased renal sympathetic efferent nerve activity and this response was blocked by cooling or sectioning the cervical vagi (Linden, Mary & Weatherill, 1980). In functional terms, atrial stretch in the dog (Prosnitz & DiBona, 1978) and the rat (Kaufman, 1984) caused an increase in sodium excretion in the absence of any change in renal blood flow. Arterial baroreceptors also lead to changes in renal sympathetic outflow as increasing the blood pressure in isolated carotid sinuses of dog (Kidd, Linden & Scott, 1981), or cat (Coote & Downman, 1969; Ninomiya & Irisawa 1975), or using pressor drugs in the rat (Coote & Sato, 1977), resulted in a powerful inhibition of renal sympathetic nerve activity. Conversely, renal sympathetic nerve activity was increased during episodes of bilateral carotid sinus occlusion (Beers, Carroll, Young & Guyton, 1986). These
reflex increases and decreases in renal nerve activity caused reciprocal changes in sodium and water excretion, with minimal alterations in renal haemodynamics (Beers et al. 1986). Together, these reports show that both the high- and low-pressure baroreceptors of the cardiovascular system can modulate sympathetic outflow to the kidneys. There is evidence from electrophysiological studies which show that afferent nerve and impulses arising from somatic sensory receptors, cardiopulmonary receptors carotid sinus baroreceptors terminate at similar sites in the central nervous system in the rabbit (Terui, Saeki & Kumada, 1987) and cat (Ciriello & Calaresu, 1977). This et al. convergence has been observed from a functional point of view by Abboud by mediated the renal vasoconstriction attenuated as volume (1981) expansion the between of the interaction little is known else sciatic nerve stimulation. Very somatic afferent input and the cardiovascular receptors in terms of the neural control of fluid handling. Therefore the primary aim of this study was to characterize the renal haemodynamic and tubular responses to activation of the afferent nerves which arise from muscle and cutaneous sensory receptors and to determine the degree to which the cardiovascular baroreceptors modify these renal responses. The approach used was to stimulate the brachial plexus in rats which had undergone either bilateral denervation of the vagi and/or the carotid sinus nerves and to compare the renal haemodynamic and functional responses with those from animals in which all cardiovascular sensory nerves were intact.
SOMATIC RECEPTORS AND RENAL FUNCTION
575
METHODS
Male albino Sprague-Dawley rats (320-400 g) were fasted overnight. They were anaesthetized with sodium pentobarbitone, 60 mg kg-' intraperitoneally, which was maintained by a 3 mg kg-' h-', intravenous infusion. The right brachial artery was cannulated to measure systemic arterial blood pressure (Statham P23 ID transducer to Hato Rey, Puerto Rico and Grass model 7 polygraph, Quincey, MA, USA), and to allow removal of blood samples. The femoral vein was cannulated to permit infusion of saline (150 mM-NaCl) and drugs. An intravenous infusion of saline, at 6 ml h-', was begun immediately after cannulation and lasted for the duration of the experiment. An iliac artery was cannulated such that the cannula tip lay in the aorta just below the level of the left renal artery. A thread was passed around the aorta, rostral to the left renal artery, and was attached to a screw device which, by loosening or tightening enabled renal arterial pressure to be regulated at control levels. The left brachial nerve plexus was isolated from surrounding tissue and placed on bipolar stimulating electrodes. The left kidney was exposed via a mid-line abdominal incision, its ureter was cannulated and the renal artery cleared to allow fitting of an electromagnetic flow probe (Carolina EP100 series probe and FM501 flowmeter, King, NC, USA). Zero blood flow was obtained by transiently occluding the renal artery. On completion of surgery a 2 ml primer (10 mg ml-' inulin in saline) was given intravenously and the infusion changed to one containing inulin, 10 mg kg-', in saline. Measurements were begun 2 h later. Experimental protocol. This consisted of five 15 min clearance periods, two before and two following a period during which the brachial nerve plexus was stimulated. A Grass model 8 stimulator (Quincey, MA, USA) provided square-wave pulses, at 15 V, 0-2 ms duration and 3 Hz frequency, which were used to stimulate the brachial nerve plexus. At least 5 min were allowed from the beginning of stimulation before the start of the urine collection for adjustment of the renal perfusion pressure to the pre-stimulus value and for pre-formed urine to clear the dead-space of the cannula. Analyses. Arterial blood samples (0-6 ml) were collected into cooled syringes at the beginning and end of the first and last pair of clearance periods. The samples were centrifuged immediately, and the plasma removed and stored at -20 00, while the cells were resuspended in an equal volume of saline and reinfused into the animal as soon as possible. Inulin was measured as previously described (Johns, Lewis & Singer, 1976) and its clearance taken as a measure of glomerular filtration rate. Plasma and urine sodium concentration was measured by emission spectroscopy (Corning 410c). Intact group. In this group all baroreceptor afferent nerves and renal sympathetic efferent nerves remained intact. Renal denervation. During the surgical preparation, all nerves arising from the coeliac ganglion were sectioned, the renal artery was painted with absolute alcohol and the denervation was confirmed by showing that the renal blanching, which occurred in response to a 5 s period of 10 Hz electrical stimulation of the coeliac ganglion, was abolished. Vagotomy. Both vagi were sectioned in the neck region as they ran parallel with the common carotid arteries. This was carried out 1 h following the administration of the primer. Carotid sinus denervation. Using a Zeiss model 212 surgical microscope, both internal carotid arteries were stripped of all nervous and connective tissue from the carotid bifurcation to the point at which they entered the skull and were then painted with absolute alcohol. This procedure was undertaken during the main surgical preparation. The combined carotid sinus denervation and vagotomy was performed during the surgical preparation. Statistics. The average value of the two clearance periods before and the two following brachial nerve stimulation (control) was compared to that obtained during the period of stimulation (experimental). The absolute and percentage values quoted represent means + standard error of the mean. Student's paired t test was carried out on data within individual groups, the difference between means being taken to be statistically significant at the 5 % level. Statistical comparisons between groups were made using a one-way analysis of variance.
G. DAVIS AND E. J. JOHNS
576
TABLE 1. Haemodynamic and renal responses to brachial nerve stimulation in the intact rat and renally denervated animals Intact (n = 6) Denervated (n = 5)
SBP (mmHg) RPP (mmHg) RBF (ml min-' kg-') GFR (ml min-' kg-') UV (1u min-' kg-') UNaV (,umol min-' kg-')
Basal 113+6 105+6
15-5+P15
Expt 137+3*** 105+6 132 + 12**
Recovery 114+6
109+7 15-0+0'6
Basal 106+3 105+3
Expt 127+4*** 106+3
16-2+P17
16-4+P15
Recovery 111+5
108+4 17-3+P14
3416+0-22 3 30+ 0-36 3'33+0 40
3-17 +±036
33-6+3#4 21-8 +36*** 32-0+3-3 84-8+13X8 83X8+14X1 7-06 +106 451 + 124*** 6-83 + 144 16-27 + 2-17 16-31 + 2'85
20-38 + 289
3'56+0-28 2-61 + 0-25*
10P8+156
FENa (%)
1-27+0-22 1.00 + 0.23* 1P31+ 0-25 3-36 +0-53 3-41+ 0-52 4-37 +0 59 The protocol consisted of five clearance periods, two before (Basal), one during (Expt) and two following (Recovery) brachial nerve stimulation. Abbreviations: SBP = systemic arterial blood pressure; RPP = renal perfusion pressure; RBF = renal blood flow; GFR = glomerular filtration rate; UV= urine flow rate; UNV = absolute sodium excretion; FENa = fractional sodium excretion. The results are displayed as mean + standard error of the mean. n = number of animals. *=P
Journal of Physiology (1991), 432, pp. 573-584 With 2 figures Printed in Great Britain
EFFECT OF SOMATIC NERVE STIMULATION ON THE KIDNEY IN INTACT, VAGOTOMIZED AND CAROTID SINUS-DENERVATED RATS
BY GERARD DAVIS AND EDWARD J. JOHNS From the Department of Physiology, The Medical School, Birmingham B15 2TT
(Received 6 March 1990) SUMMARY
1. The influence of cardiopulmonary and arterial baroreceptors on the renal nervedependent functional responses of the kidney to electrical stimulation of somatic afferent nerves was studied in pentobarbitone-anaesthetized rats. 2. Electrical stimulation of the left brachial nerve plexus at 3 Hz, 0-2 ms and 15 V in the intact animals increased blood pressure by 22 %, and while renal perfusion pressure was maintained at pre-stimulus levels, renal blood flow and glomerular filtration rate decreased by 14 and 22 % respectively. At the same time urine flow rate and absolute and fractional sodium excretion decreased by 36, 42 and 27 % respectively. In animals subjected to acute renal nerve section these renal functional responses could not be elicited. 3. Following bilateral vagotomy the systemic and renal haemodynamic responses to brachial nerve stimulation were similar to the intact group. However, urine flow rate and absolute and fractional sodium excretions decreased by 50, 59 and 47 % respectively, responses which were significantly greater than in the intact group. 4. In a group of rats in which the carotid sinus nerves had been sectioned, stimulation of the brachial plexus caused reductions of renal blood flow and glomerular filtration rate of the same magnitude as in the intact group; however, urine flow rate and absolute and fractional sodium excretion fell by 51, 60 and 48%, respectively, which were significantly larger than in the intact group. 5. These results demonstrate that the afferent nerve information arising from muscle joints and skin and carried via the brachial plexus caused reflex renal nervedependent reductions in renal haemodynamics and an antidiuresis and antinatriuresis. The cardiopulmonary and carotid sinus baroreceptors exert a tonic inhibitory action on these reflex renal responses insofar as they appeared to attenuate the antidiuretic and antinatriuretic responses to somatic afferent nerve stimulation. INTRODUCTION
The exercise pressor reflex has both peripheral and central components (Mitchell, 1985). The peripheral limb of the reflex is initiated by activation of group IV, somatic afferent nerve fibres, arising from sensory receptors in skin, joints and muscle, which discharge into group III and IV 'somatic' afferent nerves (Coote, Hilton & PerezGonzalez, 1971; Mitchell, Kaufman & Iwamoto, 1983), resulting in an increased MS 8330
574
G. DAVIS ANDE. J. JOHNS
sympathetic nerve activity, which functionally increases heart rate (Sato & Schmidt, 1987) and peripheral resistance (Thames & Abboud, 1979). With regard to the kidney, Abboud, Mark & Thames (1981) showed that brief periods of high-frequency stimulation of the somatic afferent fibres contained within the sciatic nerve induced a profound decrease in renal blood flow which was approximately frequency related. Handa & Johns (1987), using the rat, found that stimulation of the somatic sensory fibres of the brachial nerve plexus at much lower frequencies increased renal sympathetic nerve activity which was associated with only minor falls in renal haemodynamics but large decreases in the excretion of sodium. At the level of the central nervous system a number of interactions may occur. It is now clear that the pressure receptors of the cardiovascular system can influence renal sympathetic nerve activity. Karim, Kidd, Malpus & Penna (1972) demonstrated that raising left atrial pressure in the dog, by inflating a small balloon in the pulmonary vein-left atrial junction, resulted in a decreased renal sympathetic efferent nerve activity and this response was blocked by cooling or sectioning the cervical vagi (Linden, Mary & Weatherill, 1980). In functional terms, atrial stretch in the dog (Prosnitz & DiBona, 1978) and the rat (Kaufman, 1984) caused an increase in sodium excretion in the absence of any change in renal blood flow. Arterial baroreceptors also lead to changes in renal sympathetic outflow as increasing the blood pressure in isolated carotid sinuses of dog (Kidd, Linden & Scott, 1981), or cat (Coote & Downman, 1969; Ninomiya & Irisawa 1975), or using pressor drugs in the rat (Coote & Sato, 1977), resulted in a powerful inhibition of renal sympathetic nerve activity. Conversely, renal sympathetic nerve activity was increased during episodes of bilateral carotid sinus occlusion (Beers, Carroll, Young & Guyton, 1986). These
reflex increases and decreases in renal nerve activity caused reciprocal changes in sodium and water excretion, with minimal alterations in renal haemodynamics (Beers et al. 1986). Together, these reports show that both the high- and low-pressure baroreceptors of the cardiovascular system can modulate sympathetic outflow to the kidneys. There is evidence from electrophysiological studies which show that afferent nerve and impulses arising from somatic sensory receptors, cardiopulmonary receptors carotid sinus baroreceptors terminate at similar sites in the central nervous system in the rabbit (Terui, Saeki & Kumada, 1987) and cat (Ciriello & Calaresu, 1977). This et al. convergence has been observed from a functional point of view by Abboud by mediated the renal vasoconstriction attenuated as volume (1981) expansion the between of the interaction little is known else sciatic nerve stimulation. Very somatic afferent input and the cardiovascular receptors in terms of the neural control of fluid handling. Therefore the primary aim of this study was to characterize the renal haemodynamic and tubular responses to activation of the afferent nerves which arise from muscle and cutaneous sensory receptors and to determine the degree to which the cardiovascular baroreceptors modify these renal responses. The approach used was to stimulate the brachial plexus in rats which had undergone either bilateral denervation of the vagi and/or the carotid sinus nerves and to compare the renal haemodynamic and functional responses with those from animals in which all cardiovascular sensory nerves were intact.
SOMATIC RECEPTORS AND RENAL FUNCTION
575
METHODS
Male albino Sprague-Dawley rats (320-400 g) were fasted overnight. They were anaesthetized with sodium pentobarbitone, 60 mg kg-' intraperitoneally, which was maintained by a 3 mg kg-' h-', intravenous infusion. The right brachial artery was cannulated to measure systemic arterial blood pressure (Statham P23 ID transducer to Hato Rey, Puerto Rico and Grass model 7 polygraph, Quincey, MA, USA), and to allow removal of blood samples. The femoral vein was cannulated to permit infusion of saline (150 mM-NaCl) and drugs. An intravenous infusion of saline, at 6 ml h-', was begun immediately after cannulation and lasted for the duration of the experiment. An iliac artery was cannulated such that the cannula tip lay in the aorta just below the level of the left renal artery. A thread was passed around the aorta, rostral to the left renal artery, and was attached to a screw device which, by loosening or tightening enabled renal arterial pressure to be regulated at control levels. The left brachial nerve plexus was isolated from surrounding tissue and placed on bipolar stimulating electrodes. The left kidney was exposed via a mid-line abdominal incision, its ureter was cannulated and the renal artery cleared to allow fitting of an electromagnetic flow probe (Carolina EP100 series probe and FM501 flowmeter, King, NC, USA). Zero blood flow was obtained by transiently occluding the renal artery. On completion of surgery a 2 ml primer (10 mg ml-' inulin in saline) was given intravenously and the infusion changed to one containing inulin, 10 mg kg-', in saline. Measurements were begun 2 h later. Experimental protocol. This consisted of five 15 min clearance periods, two before and two following a period during which the brachial nerve plexus was stimulated. A Grass model 8 stimulator (Quincey, MA, USA) provided square-wave pulses, at 15 V, 0-2 ms duration and 3 Hz frequency, which were used to stimulate the brachial nerve plexus. At least 5 min were allowed from the beginning of stimulation before the start of the urine collection for adjustment of the renal perfusion pressure to the pre-stimulus value and for pre-formed urine to clear the dead-space of the cannula. Analyses. Arterial blood samples (0-6 ml) were collected into cooled syringes at the beginning and end of the first and last pair of clearance periods. The samples were centrifuged immediately, and the plasma removed and stored at -20 00, while the cells were resuspended in an equal volume of saline and reinfused into the animal as soon as possible. Inulin was measured as previously described (Johns, Lewis & Singer, 1976) and its clearance taken as a measure of glomerular filtration rate. Plasma and urine sodium concentration was measured by emission spectroscopy (Corning 410c). Intact group. In this group all baroreceptor afferent nerves and renal sympathetic efferent nerves remained intact. Renal denervation. During the surgical preparation, all nerves arising from the coeliac ganglion were sectioned, the renal artery was painted with absolute alcohol and the denervation was confirmed by showing that the renal blanching, which occurred in response to a 5 s period of 10 Hz electrical stimulation of the coeliac ganglion, was abolished. Vagotomy. Both vagi were sectioned in the neck region as they ran parallel with the common carotid arteries. This was carried out 1 h following the administration of the primer. Carotid sinus denervation. Using a Zeiss model 212 surgical microscope, both internal carotid arteries were stripped of all nervous and connective tissue from the carotid bifurcation to the point at which they entered the skull and were then painted with absolute alcohol. This procedure was undertaken during the main surgical preparation. The combined carotid sinus denervation and vagotomy was performed during the surgical preparation. Statistics. The average value of the two clearance periods before and the two following brachial nerve stimulation (control) was compared to that obtained during the period of stimulation (experimental). The absolute and percentage values quoted represent means + standard error of the mean. Student's paired t test was carried out on data within individual groups, the difference between means being taken to be statistically significant at the 5 % level. Statistical comparisons between groups were made using a one-way analysis of variance.
G. DAVIS AND E. J. JOHNS
576
TABLE 1. Haemodynamic and renal responses to brachial nerve stimulation in the intact rat and renally denervated animals Intact (n = 6) Denervated (n = 5)
SBP (mmHg) RPP (mmHg) RBF (ml min-' kg-') GFR (ml min-' kg-') UV (1u min-' kg-') UNaV (,umol min-' kg-')
Basal 113+6 105+6
15-5+P15
Expt 137+3*** 105+6 132 + 12**
Recovery 114+6
109+7 15-0+0'6
Basal 106+3 105+3
Expt 127+4*** 106+3
16-2+P17
16-4+P15
Recovery 111+5
108+4 17-3+P14
3416+0-22 3 30+ 0-36 3'33+0 40
3-17 +±036
33-6+3#4 21-8 +36*** 32-0+3-3 84-8+13X8 83X8+14X1 7-06 +106 451 + 124*** 6-83 + 144 16-27 + 2-17 16-31 + 2'85
20-38 + 289
3'56+0-28 2-61 + 0-25*
10P8+156
FENa (%)
1-27+0-22 1.00 + 0.23* 1P31+ 0-25 3-36 +0-53 3-41+ 0-52 4-37 +0 59 The protocol consisted of five clearance periods, two before (Basal), one during (Expt) and two following (Recovery) brachial nerve stimulation. Abbreviations: SBP = systemic arterial blood pressure; RPP = renal perfusion pressure; RBF = renal blood flow; GFR = glomerular filtration rate; UV= urine flow rate; UNV = absolute sodium excretion; FENa = fractional sodium excretion. The results are displayed as mean + standard error of the mean. n = number of animals. *=P
