Brain Research, 522 (1990) 99-106

99

Elsevier BRES 15646

Cardiovascular responses to electrical stimulation of the ventrolateral medulla of the spontaneously hypertensive rat R.K.W. Chan, Y.S. Chan and T.M. Wong Department of Physiology, Facultyof Medicine, Universityof Hong Kong (Hong Kong) (Accepted 2 January 1990)

Key words:Rostral ventrolateral medulla; Blood pressure; Heart rate; Responsiveness; Electrical stimulation; Spontaneously hypertensive rat; Wistar-Kyoto rat

Cardiovascular responses to electrical microstimulation of the ventrolateral medulla were investigated in both Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) under pentobarbitai anesthesia. The threshold intensity required to elicit a change in blood pressure (BP) and the cardiovascular responses in these two groups of rats upon electrical stimulation were compared. It was found that the region with the lowest threshold intensities was located in the rostral ventrolateral medulla (RVL) and the threshold intensities were much lower in SHR than in WKY. Electrical stimulation of this brain region also resulted in a greater increase in BP during stimulation in SHR, compared to control. In SHR, upon termination of stimulation, the BP dropped to a level above the pre-stimulation level and this was followed by a prolonged, sustained elevation in BP before returning to the control level, whereas in WKY, the BP showed an initial drop to below the pre-stimulation level and then returned to the control level. These results suggest an enhanced responsiveness to electrical stimulation in SHR. Although the heart rate (HR) increased to a similar extent during electrical stimulation in both groups of rats, upon termination of stimulation WKY exhibited bradycardia followed by tachycardia before the HR returned to the pre-stimulation level, whereas SHR exhibited tachycardia which was maintained for a substantial period of time before returning to the pre-stimulation level. The results suggest a lower baroreceptor sensitivity in SHR. The change in HR/change in BP was smaller in SHR than in WKY, suggesting that the increase in HR may not contribute to the increase in BP during stimulation as much in SHR as in WKY. The results confirm that RVL is important in the control of circulation. The lower threshold in eliciting cardiovascular changes, greater responsiveness to electrical stimulation and lower baroreceptor sensitivity may be responsible for the development of hypertension in SHR. INTRODUCTION It has b e e n shown that application of inhibitory substances which block synaptic transmission in the ventral m e d u l l a p r o d u c e d a great fall in b l o o d pressure s'9, w h e r e a s excitatory substances had the opposite effect 7, suggesting that this a r e a m a y be a tonic centre in the control of circulation. The finding was subsequently c o n f i r m e d and substantiated. In addition to being a tonic centre, the ventral m e d u l l a m a y also be crucial in the reflex control of circulation as it was found to be linked functionally with brain regions known to be involved in the control of circulation, notably the nucleus tractus solitarius 4"6'21'32, the p e r i a q u e d u c t a l grey and defence a r e a of the h y p o t h a l a m u s 1°'14'34, and the fastigial nucleus 2° which has been shown to m o d u l a t e the cardiovascular responses during exercise 5. N e u r o n s within the ventral m e d u l l a have been shown to project to the i n t e r m e d i o l a t e r a l column of the spinal cord where the preganglionic a u t o n o m i c fibres arise 1'2'4'16'19'23'3°. In short, ventral m e d u l l a m a y be a generating centre for s y m p a t h e t i c outflow as well as a relay station of various

reflexes involved in the control of circulation. Thus, abnormalities in this a r e a m a y lead to alterations in sympathetic outflow, and eventually abnormalities in cardiovascular functions. It has b e e n d e m o n s t r a t e d by direct m e a s u r e m e n t of s y m p a t h e t i c nerve activity in anesthetized 13 o r conscious 18'34 s p o n t a n e o u s l y hypertensive rats ( S H R ) , an i n b r e d strain o f the W i s t a r - K y o t o rat ( W K Y ) which develops h y p e r t e n s i o n s p o n t a n e o u s l y , that the sympathetic outflow is increased, suggesting that an increased s y m p a t h e t i c activity m a y contribute to hypertension in these animals. In s u p p o r t of this, it has also been found that b o r d e r l i n e hypertensive rats develop sustained h y p e r t e n s i o n a c c o m p a n i e d by an increased capacity for biosynthesis, storage and release of catecholamines in the adrenal m e d u l l a in response to chronic stress 22. The results of this study further suggest that the response in b l o o d pressure of the b o r d e r l i n e hypertensive rat is g r e a t e r in chronic stress, which m a y be related to the increased sympathetic activity22. Thus it is likely that the e n h a n c e d cardiovascular responses and sympathetic activity in the b o r d e r l i n e hypertensive rat m a y result from altered activities in the ventral medulla. To test this

Correspondence: T.M. Wong, Department of Physiology, Faculty of Medicine, University of Hong Kong, 5 Sassoon Road, Hong Kong. 0006-8993/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)

100

hypothesis, we stimulated the ventrolateral medulla electrically and compared the responses in arterial blood pressure (BP) and heart rate (HR) between the normotensive WKY and the borderline hypertensive rats of the SHR strain. Part of this study has been presented as an abstract3.

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MATERIALS AND METHODS Female rats of WKY (n = 18) and SHR (n = 22) of 210-230 g (19-20 weeks old) were used. They were anesthetised with pentobarbital sodium (Sagatai, May and Baker Ltd.) at an initial dose of 40 mg/kg body weight (i.p.), supplemented by 10 mg/kg

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Fig. 1. A: diagram of the medulla depicting sites of electrical stimulation (40/~A) along electrode tracks a, b and c and the associated blood pressure changes in SHR. Tracks a and c are 1.6 mm from the midline while track b is 2.1 mm from the midline. Tracks a and b are located at 12.5 mm posterior to bregma while tracks c is located at 13.3 mm posterior to bregma. The dotted circle represents the estimated current spread at 40/~A. The dotted lines under the bottom 3 tracings on the left panel and the bottom 2 tracings on the right panel are drawn for reference. The thick horizontal bars at the bottom of the left, middle and right panels indicate the duration of electrical stimulation. B: diagram of serial sections through the medulla showing a summary of all the stimulation sites from 22 SHR rats and the associated blood pressure changes. The vertical lines depict the location of the stimulation tracks, which are separated by 200 #m. Filled circles represent areas having pressor and post-stimulation pressor responses; open circles represent areas having pressor responses only; inverted triangles represent areas having depressor responses. NTS, nucleus tractus solitarius; AMB, nucleus ambiguus; RVL, rostral ventrolateral medulla; PGL, paragigantocellularis lateralis nucleus; LRN, lateral reticular nucleus.

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(i.v.) as required. All pressure points and wound edges were infiltrated with lidocaine. The animals respired spontaneously; the respiratory rate was not significantly disturbed by any of the manipulations employed in this study. A femoral artery was cannulated with a polyvinylethylene tubing connected to a Statham pressure transducer and Gould recorder for the measurement of BP and HR. The head of the rat was then mounted to a stereotaxic apparatus (Narishige, Japan) and was maintained without any apparent signs of pain. The occipital bone overlying the cerebellum was surgically removed to allow placement of the stimulating microelectrodes. An array of two parallel tungsten microelectrodes (tip diameter size 1/~m, 10 MK2, Frederick Hear) separated by 0.2 mm was inserted vertically through the intact cerebellum in a dorsoventral fashion to the ventrolateral medulla on the right side (according to the rat's brain atlas 25) for monopolar cathodal microstimulation. The ventrolateral medulla was systematically explored with an electrical current of 40 /~A, starting with the microelectrode 4 mm from the ventral surface of the medulla. Sites along each electrode track were investigated in steps of 200/tm until the tip of the microelectrode was at an estimated depth of 0.5 mm from the ventral surface of the medulla. For systemic mapping, a territory with coordinates 10.8-13.3 mm caudal from the bregma and 1.5-2.3 mm lateral from the midline was explored, using tracks separated by 0.2 mm in both rostrocaudal and mediolateral directions. About 16-18 electrode penetrations were made in each rat. Electrical stimulation was delivered from a Grass $48 stimulator via a stimulation isolation unit (Grass SIU5A) and a constantcurrent unit (Grass CCU1A). At each stimulation site, a train of rectangular pulses of 2 ms duration at 70 Hz was passed for 5 s. Peak-to-peak current was monitored throughout the stimulation period as the voltage drop across a 1 kt2 resistor, and was displayed on a storage oscilloscope. The current used in the present study ranged from 0.5-100/~A. BP and HR were monitored continuously throughout the experiment. The rectal temperature of the animal was maintained at 37 °C by a thermostatically controlled heating device. At each stimulation site within the vicinity of the rostral ventrolateral medulla, the threshold current was determined. The threshold current was arbitrarily defined as the amount of current required to elicit a change of 2.5 mmHg in BP (as compared with the prestimulation value) which was the highest resolution of the recording system used. Routinely, electric current of 40 pA was first applied to determine if any cardiovascular changes could be evoked. If responses could be evoked with this current, the current density was gradually decreased until 0.5 # A for determination of the threshold current. If no cardiovascular responses were evoked with 40 #A, the current density was increased progressively until 100 #A.

After the threshold current was determined at each test site, currents of different intensities (10, 20, 40, 70 gA) were delivered in random sequence at each location and the changes in BP and HR were recorded. For each animal studied, about 6-9 small electrolytic lesions (0.5 mA DC for 5 s) were made at the most ventral site of electrode penetrations of at least 0.4 mm apart. In addition, at the final stage of each experiment, one lesion was made at the site where the most prominent cardiovascular responses could he evoked. At the end of each experiment, the brain was peffused with 10% formal saline solution. The brain was frozen and frontal sections of 60 #m were cut with a cryostat. The stimulation sites were reconstructed on cresyl violet stained serial sections with reference to coordinates of the electrode tracks and electrolytic lesions. Student's t-test was used to analyse the difference in change in BP upon electrical stimulation between the two groups of rats. Regression lines showing the relationship between change in HR and change in BP in both rats were drawn. Test for parallelism of the regression lines of the two groups of rats was made. A difference at the level of P < 0.05 was considered statistically significant.

RESULTS T h e first series o f e x p e r i m e n t s o f t h e p r e s e n t s t u d y was to c o m p a r e t h e c h a n g e s in B P u p o n e l e c t r i c a l s t i m u l a t i o n of d i f f e r e n t sites in t h e v e n t r o l a t e r a l m e d u l l a b e t w e e n W K Y a n d S H R . Fig. 1 s h o w s t h e d i s t r i b u t i o n o f t h e B P r e s p o n s e p a t t e r n s u p o n s t i m u l a t i o n o f d i f f e r e n t sites o f the v e n t r o l a t e r a l m e d u l l a in S H R . T h r e e c h a r a c t e r i s t i c B P r e s p o n s e s w e r e e v o k e d in d i f f e r e n t r e g i o n s within the t e r r i t o r y e x p l o r e d with s t i m u l a t i o n c u r r e n t o f 1 0 - 7 0 / t A in e i t h e r W K Y o r S H R ; t w o r e s p o n s e types w e r e similar while one

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