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Responses of lumbar vasoconstrictor neurons supplying different vascular beds to graded baroreceptor stimuli in the cat
Journal of the Autonomic Nervous System, 42 tia93) 241-250 © 1993 Elsevier Science Publishers B.V. All rights reserved 0165-1838/93/$06.00
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JANS 01369
Responses of lumbar vasoconstrictor neurons supplying different vascular beds to graded baroreceptor stimuli in the cat M. M i c h a e l i s , A . B o c z e k - F u n c k e , H . - J . H h b l e r a n d W . J[inig Physiologisches Ir..stitut, Christian-Albrechts-Universitiit, Kiel, FRG
(Received 4 August 1992) (Reviston received 14 October 1992) (Accepted 14 October 1992)
K ~ , words': Sympathetic vasoconstrictor neurons; B a r o r e c e p t o r reflex; Cat
Abstract Lumbar ~ympathetic vasoconstrictor neurons supplying skeletal muscle, hai~ skLn. and pelvic organs were tested for their responses to carotid haroreceptor stimulation in ehloralose-anaesthetized cats. Us:ng single- and few-fibre recordings, the responses of the different types of vasoconstrictor neuron to graded steps of non-pulsati;e pressure ranging from 110 to 260 mmHg in a vascularly isolated carotid sinus were analyzed quantitatively during the first 10 s of stimulation. The activity in all postganglionic muscle vasoconstrictor (MVC) neurons, preganglionic visceral vasoconstrictor (VVC) neurons and one third of the poszganglionic cutaneous vasoconstrictor (CVC 1) neurons was strongly depressed by maximal baroreceptor stimulation. Moreover, quantitative analysis revealed no significant differences of the baroreceptor sensitivity of MVC and CVC,. neurons as compared with VVC neurons at all levels of carotid sinus pressure. In contrast, two-thirds of the postganglionic cutaneous vasoconstrictor (CVC 2) neurons exhibited a significantly weaker barosensitivity. The functional implications are discussed.
Introduction T h e s p o n t a n e o u s activity in many pre- a n d postganglionic sympathetic n e u r o n s is u n d e r inhibitory control of arterial baroreceptors. However, d e t e c t a b l e barosensitivity is not a c o m m o n f e a t u r e o f all sympathetic neurons. Thus, the activity in only about 25% o f the preganglionic sympathetic ne'.:,ro~s projecting into the l u m b a r splanchnic nerves is inhibited by b a r o r e c e p t o r stimulation during the systolic rise of the pulse p r e s s u r e wave or large pressure increases in an isolated carotid blind sac [2,3]. On the o t h e r
Correspondence to: M. Michaelis, Physiologisches Institut, Christian-AIbrechts-Universit~it, OIshausenstr. 40, D-2300 Kiel, FRG.
hand, all postganglionic sympathetic neurons with s p o n t a n e o u s activity projecting to skeletal muscle of the hindlimb show strong barosensitivity. These neurons very likely form a functionally homogeneous group of muscle vasoconstrictor (MVC) neurons [4]. Although the postganglionic sympathetic fibres projecting to hairy skin of the hindlimb, which are spontaneously active, also have vasoconstrictor function, only roughly onethird of these neurons exhibits a clearly detectablc barosensitivity [4,19,20]. Similar differences b e t w e e n M V C and cutaneous vasoconstrictor (CVC) n e u r o n s regarding their barosensitivity exist in h u m a n muscle and skin nerves [14,15,31]. N u m e r o u s studies revealed quantitatively diff e r e n t responses of specific vascular beds to baror e c e p t o r stimulation [7,9,13,16,17,22,33]. However, these studies give only indirect information
242 about the neuronal components of these reflex responses. Comparing whole nerve recordings, Ninomiya and coworkers [27,28] described non-uniform responses to baroreceptor stimulation in sympathetic fibres to heart, kidney, spleen and intestine. On one hand this could be due to quantitative differences in the barosensitivity of a uniform population of vasoconstrictor neurons projecting to the various organs. On the other, the recorded nerves could contain different proportions of vasoconstrictor neurons of quantitatively similar barosensitivity. In the present experiments we recorded activity from single- or few-fibre preparations of MVC, CVC and visceral vasoconstrictor neurons and quantitatively studied the responses of these neurons to graded stimulation of the arterial baroreceptors. Some of the results have been published in preliminary form [5].
Materials and Methods
Anaesthesia and animal maintenance Experiments were performed on 12 cats of either sex weighing 2.5-4.7 kg. Anaesthesia was induced by injection of ketamine (Ketanest R, Parke-Davis, 15-20 m g / k g i.m.) and continued with a-o-glucochloralose (a-Chloralose R, Serva, 40 m g / k g i.p.). Supplementary doses of a-D-glucochioralose were given intravenously (10 m g / k g ) to maintain deep anaesthesia as judged from miotic pupils and absence of spontaneous blood pressure and heart rate fluctuations. Systemic arterial blood pressure was continuously recorded using a pressure transducer (List LM-22) connected to a cannula in the femoral or brachial artery. Throughout the experiments mean arterial blood pressure exceeded 90 mmHg. The jugular vein was cannulated for drug administration. Animals were immobilized with pancuronium bromide (Pancuronium-Organon R, Organon Teknika, 0.8 m g / h i.v.) and artificially ventilated through a tracheal cannula keeping the end-expiratory CO z at 3.5-4.5 vol%. Body core temperature was recorded intra-oesophageally and
maintained at 37-38°C by a heating plate. The urinary bladder was catheterized transurethrally to monitor urine excretion. At the end of the experiments the animals were killed by int~avenous injection of a saturated potassium chloride solution under deep anaesthesia. All experiments had been approved by the local animal care committee of the state administration and were conducted in accordance with the German federal law.
Surgery The region of the left carotid sinus was vascularly isolated by ligating the internal carotid artery and the large branches of the external carotid artery. A cannula inserted into the left external carotid artery was connected to a pressure reservoir filled with saline at 25°C and to a pressure transducer (List LM-22) in order to measure the carotid sinus pressure continuously° In all experiments the right carotid sinus nerve was cut. The aortic nerves were left intact. In six experiments the !Left sympathetic trunk and lumbar spianchnic nerves were exposed using a lateral approach, One lumbar splanchnic nerve was dissected free from surrounding tissue and mounted on a perspex platform. Subsequently, exposed tissue and nerves were covered with warm paraffin oil in a pool made out of skin flaps. In a second set of six experiments the deep and superficial peroneal nerves of the left hindlimb were exposed, mounted on perspex platforms and covered with pa~'affin oil. Nerve strands were isolated fiom the lumbar splanchnic nerves, cut peripherally and split until single unit activity could be recorded. Strands containing several postganglionic axons with spontaneous activity were dissected from those parts of the deep peroneal nerve which innervate the anterior tibial muscle and the extensor digitorum longus muscle and from the main part of the superficial peroneal nerve which innervates hairy skin.
Recording Neuronal activity was recorded unipolarly with respect to a reference electrode which was attached to the nearby tissue and was amplified by
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a low-noise AC preamplifier (input impedance 10 MI~) and a main amplifier with low input impedance, filtered with a band width of 80-120 Hz to 1000-1200 Hz.
before the onset of the stimulus (M) and mean arterial blood pressure 18 s after the onset (N):
Baroreceptor stimulation
For statistical comparison of mean decreases in the activity of different types of vasoconstrictor neurons Student's t-test was applied. Average responses of vasoconstrictor neurons were tested for significance with the paired t-test.
The carotid blind sac could be closed reversibly by a snare which was set around the commc, n carotid artery. As a result the carotid sinus pressure in the closed blind sac fell to 3 0 - ! 0 0 mmHg whilst the systemic blood pressure markedly increased (see Fig. 2B). This residual non-pulsatile pressure was presumably due to retrograde blood flow through small untied arteries. The carotid baroreceptors were stimulated by stepwise raising the non-pulsatile pressure in the closed blind sac to 110-260 mmHg keeping the pressure level constant for at least 10 s. Between two stimuli the carotid sinus pressure was lowered to the residual level. The stimulation protocol usually consisted of a series of five different pressure steps with differences of 30 mmHg, starting with a stimulus in the range of 110-140 mmHg up to a stimulus in the range of 230-260 mmHg.
Data analysis Neuronal activity, carotid sinus pressure, mean arterial blood pressure and ECG were stored on magnetic tape (EMI SE 7000). During .off-line analysis, neuronal activity was fed through window discriminators to produce standard impulses. The decrease in neuronal activity and the reduction of mean arterial blood pressure induced by stimulation of carotid baroreceptors were quantified using a computer program. The relative decrease in activity ( R D A ) was calculated from the relation between the summed neuronal activity during 10 s before the onset of the stimulus (A) and the summed neuronal activity of the first 10 s after onset of the stimulus (B): R D A = ( A - B ) A - t 100%. The relative decrease in blood pressure ( R D B ) was calculated from the relation between the mean arterial blood pressure during the 10 s
RDB=(M-N)M-IIO0%.
Results
Functional identification of vasoconstrictor neurOllS
We recorded activity from eight filaments containing few (3-7) postganglionic units with spontaneous activity projecting to skeletal muscle of the hindlimb. In all cases the activity showed a strong ECG-related rhythmic modulation (cardiac rhythmicity). These neurons will be referred to as muscle vasoconstrictor (MVC) neurons. Activity in postganglionic sympathetic neurons projecting to hairy skin of the hindlimb was recorded from thirteen nerve strands (nine fewfibre preparations, four single fibres). These neurons were tentatively classified as cutaneous vasoconstrictor (CVC) neurons for two reasons: vasodilator neurons to hairy skin exhibit no spontaneous activity and pilomotor and sudomotor neurons innervating the hairy skin of the hindlimb most likely do not exist (see [19]). The activity in four out of 13 CVC preparations showed a strong cardiac rhythmicity, whereas no or weak pulse synchronous modulation could be detected in the activity of the remaining CVC neurons. This proportion is consistent with the proportion of CVC neurons showing strong cardiac rhythmicity reported in a previous study [4]. Preganglionic neurons projecting in the lumbar splanchnic nerves were classified as visceral vasoconstrictor (VVC) neurons when they were under clear inhibitory control of the arterial baroreceptors (see [3]). In the present study the activity in seven VVC neurons was analysed. Figure IA shows a typical example. The activity in this VVC neuron exhibited a strong cardiac rhythmicity and
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was d e p r e s s e d during a high pressure stimulus in the carotid sinus blind sac (Fig. 1A). T h e majority of preganglionic n e u r o n s projecting in the lumbar splanchnic nerves did not res p o n d to stimulation o f the arterial b a r o r e c e p t o r s (Fig. 1B). T h e s e n e u r o n s are p r o b a b l y involved in regulation of colon and pelvic o r g a n motility a n d are t h e r e f o r e called motility-regulating ( M R ) n e u r o n s [2].
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2rain Fig. 2 Responses of a single preganglionie visceral vasoconstrictor neuron (VVC) to graded isotonic; pressure stimuli applied to the carotid baroreceptors. (A): response to maximal baroreceptor stimulus. Traces from above: neuronal activity, carotid sinus pressure (CSP) and blood pressure (BP). Regular blood pressure oscillations are induced by artificial ventik:tion. (B): neuronal activity given as a peristimulus-histogram, bin width 2 s (VVC). After closure of the carotid blind sac (downward arrow) CSP dropped simultaneously with the excitation of the unit while the mean arterial blood pressure (MAP) rose. Opening of the carotid blind sac (upward arrow) led to a brief marked inhibition of the unit and to a decline of MAP to resting level, Note the graded inhibition of neuronal activity depending on the magnitude of the baroreceptor stimulus. The pressure stimulus marked with an asterisk is shown as original record in A. (C): post-R-wave histogram (bin width 8 ms), 500 cycles superimposed.
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Fig. 1. Responses of a single preganglionic visceral vasoconstrictor neuron (A), a single preganglionic motility regulating neuron (B) and blood pressure to isotonic pressure stimuli applied to the carotid sinus baroreceptors. Traces from above show carotid sinus pressure (CSP), mean arterial blood pressure (MAP) and neuronal activity in the visceral vasoconstrictor neuron ( W C ) and the motility regulating neuron (MR; peristimulus-histograms, bin width 2 s). Insets show the discriminated actio~ potentials of the units, five times superimposed. The post-R-wave histograms (bin width 8 ms) were obtained from 500 superimposed sweeps over a period of two cardiac cycles.
plied to t h e carotid sinus blind sac w e r e d e t e r mined. As o u t l i n e d in M a t e r i a l s a n d M e t h o d s the residual i n t r a c a r o t i d p r e s s u r e r a n g e d b e t w e e n 30 and 100 m m H g . T h e m e a n residual values did not differ b e t w e e n the two e x p e r i m e n t a l series: 50.3 + 15.9 m m H g during M V C / C V C recordings a n d 60.0 + 11.9 m m H g during V V C recordings ( P > 0.2). A typical e x p e r i m e n t is shown in Fig. 2. Each r a p i d rise of carotid sinus p r e s s u r e was followed by a p r o m p t d e c r e a s e in the activity o f
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Fig. 3. Quantified decreases in the activity of muscle (MVC), visceral (VVC) and cutaneous (CVCI, CVC 2) vasoconstrictor neurons to graded pressure steps applied to the isolated carotid sinus. Dots represent the individual data. All MVC, VVC. CVC z ~nd four of nine CVC 2 preparations were tested with five different baroreceplor stimuli each. Five CVC 2 were only tested with raaximal baroreceptor stimuli > 230 mmltg. CSP, carotid sinus pressure.
the single VVC neuron and by a concomitant fall of blood pressure (Fig. 2B). Both were dependent on the magnitude of the stimulus. Neuronal responses and induced blood pressure changes were quantified (see Materials and Methods). Maximal baroreceptor stimuli (carotid sinus pressure above 230 mmHg) depressed almost totally the activity, in all MVC and VVC neurons (Table 1), whereas CVC neurons did not respond homogeneously to this stimulus. The activity in four out of 13 CVC preparations decreased by 66% or more (CVCt; Table I). These CVC~ neurons also showed a strong cardiac rhythmicity in their a&ivity. Compared with the responses of the remaining CVC neurons (CVCz), the decreases in the activity ia CVC~ neurons as well as those in MVC and VVC neurons were significantly stronger (t-test, P < 0.001; Table I).
MVC, VVC and CVC t neurons 6xhibited no significant differences in their responses to maximal baroreceptor stimulation. Figure 3 illustrates the quantitative responses separately for the different types of vasoconstricTABLE I Ra . . . . . . . . ~f ,4ifferp,,t h ..... o f . . . . . . . . . . ;. . . . neurons to maxim a l stimulation o f carotid baroreceptors
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tor neuron. All MVC, and CVC t and four out o f nine C V C 2 p r e p a r a t i o n s were tested with five graded isotonic pressure stimuli. Five CVC preparations only tested with maximal b a r o r e c e p tor Stimuli decreased in their activity by less than 50% and were t h e r e f o r e included in the g r o u p of C V C 2 neurons. T h e responses of M V C neurons to carotid sinus pressure stimuli exceeding 140 m m H g w e r e significant relative to control activity (paired ttest, P < 0 . 0 5 ) . Carotid pressures of 110-140 m m H g induced no significant responses. Like M V C neurons, W C and CVC~ neurons exhibited significant decreases in their activity to
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carotid sinus pressures o f 140-170 m m H g and more. In contrast, CVC 2 n e u r o n s showed n o significant responses except to maximal b a r o r e c e p t o r stimuli (230-260 m m H g ) . Figure 4 depicts the a v e r a g e d data. F o r the different types o f vasoconstrictor n e u r o n the m e a n decreases in activity were calculated separately for five 3 0 - m m H g carotid sinus pressure intervals beginning with the 110-140 m m H g interval. T h e d e c r e a s e s in m e a n arterial b l o o d pressure w e r e calculated in the same way. Statistical analysis revealed no significant d i f f e r e n c e s b e t w e e n the responses in MVC, V V C a n d CVC~ neurons. However, the b a r o r e c e p t o r - i n d u c e d decreases in these t h r e e types of vasoconstrictor n e u r o n w e r e significantly g r e a t e r than those in C V C 2 n e u r o n s regarding carotid sinus pressures exceeding 170 m m H g (t-test, P < 0.05). T h e d e c r e a s e s in m e a n arterial b l o o d pressure i n d u c e d by the same stimuli in e x p e r i m e n t s in which n e u r o n a l activity was r e c o r d e d f r o m V V C alone, and simultaneously from M V C and CVC 1, and f r o m M V C and C V C 2, respectively, did not differ significantly.
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Fig. 4. Mean decreases in neuronal activity, obtained from different types of vasoconstrictor neuron, and concomitant mean decreases in mean arterial blood pressure (MAP) induced by graded isotonic baroreceptor stimuli (CSP, carotid sinus pressure). Individual data were averaged from intervals of 3U mmi-ig each. Vertical bars give 1 S.E.M. Mean responses in MVC neurons (®), neurons (#) and CVC l neurons ( I ) d i d not differ significantly from each other (t-test, P > 0.05). The mean decreases in activity in these three types of vasoconstrictor neurons were significantly greater than those in CVCe neurons (D) regarding carotid sinus pressures in excess of 170 mmHg (t-test, P < 0.05). Activity in MVC neurons was always recorded simultaneously with activity in CVC~ or CVCz neurons. Therefore, only the mean decreases in MAP obtained during recording from CVC~ (m) ana CVC 2 (12) neurons were given. Mean decreases in MAP during recording from CVC~, CVC 2 and VVC neurons ( q ) were not significantlydifferent.
WC
In the p r e s e n t study we described the quantitative responses of different types o f vasoconstrictor n e u r o n to b a r o r e c e p t o r stimulation using an isolated carotid blind sac p r e p a r a t i o n . Most likely the o b s e r v e d changes in sympathetic activity during increased carotid sinus pressures were i n d e e d due to b a r o r e c e p t o r stimulation and not due to a c o n c o m i t a n t c h e m o r e c e p t o r activation. It is known that the activity in C V C n e u r o n s without cardiac rhythmicity is inhibited by c h e m o r e c e p t o r stimulation [4]. In our study the C V C 2 n e u r o n s which s h o w e d no cardiac rhythmicity were not or only weakly inhibited by p r e s s u r e steps in the carotid blind sac. Thus, if these pressure steps would lead to a significant c h e m o r e c e p t o r activation, o n e would expect a m o r e p r o m i n e n t inhibitioJa in C V C 2 neurons. F u r t h e r m o r e , M V C neurons are strongly activated by c h e m o r e c e p t o r stimulation [4] but we o b s e r v e d an almost complete d e p r e s s i o n in their activity during large distension o f the isolated carotid sinus.
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Significant responses of MVC, VVC and C V C l neurons were detected using carotid sinus pressures of more than 140 mmHg. This relatively high intensity may have several reasons. First, compared with synchronous stimulation of carotid baroreceptor afferents on both sides, pressure steps in an unilateral carotid blind sac induce only half as large decreases in systemic blood pressure [29]. The same applies probably to the induced decreases in vasoconstrictor activity. Secondly, we left the aortic nerves intact. Thus, unloading of aortic baroreceptor affe rents during the decrease of systolic blood pressure may have partly counteracted the reflex inhibition of vasoconstrictor neurons during carotid sinus pressure stimuli. There is strong evidence that bulbospinal neurons located in the rostral ventrolateral medulla are important for the control of the activity in vasoconstrictor neurons. Furthermore, there seem to be several subpopulations of 'presympathetic' neurons which may be associated with the regulation of different vascular beds [8,23,25,26]. Thus, the differences and similarities of baroreceptor sensitivity found in different types of sympathetic vasoconstrictor neurons may reflect the specific medullary organization of presympathetic neurons. It is known from anatc,mical investigations that veins in the skeletal ml~scle, in contrast to arteries, receive no significant sympathetic innervation [10,24]. Thus, MVC neurons presumably innervate almost exclusively resistance vessels. MVC neurons are under strong inhibitory control of arterial barorecaptors as shown in this and other investigations [4,20]. It is likely that strong baroreceptor sensitivity is not only typical for MVC neurons but is a common feature of postganglionic sympathetic neurons innervating resistance vessels. Thus, CVC~ neurons which showed the same barosensitivity as MVC neurons may innervate the resistance vessels in the hairy skin. This would be consistent with results of DiSalvo et al. [9]. Measuring "the changes of resistance they observed no differences between dilatation of small muscle and skin arteries during baroreceptor stimulation. Vasoconstriction of peripheral superficial but not of proximal and deep
veins of the hindlimb could be observed during electrical stimulation of the lumbar sympathetic trunk [17,32]. However, baroreceptor stimulatior~ does not induce a significant diameter change of peripheral cutaneous veins [7,17] indicating that cutaneous vasoconstrictor neurons supplying these veins are not under significant baroreceptor control. Therefore, CVC 2 neurons may innervate peripheral cutaneous veins. Furthermore, CVC 2 neurons may play a role in thermoregulatien [i i]. VVC neurons may project to resistance as well as capacitance vessels in the visceral domain. Stimulation of arterial baroreceptors leads to a clear reflex change in abdominal vascular resistance and capacitance [16,22]. Direct observations of blood vessels during baroreceptor stiraulation have revealed significant diameter-changes in both arterial and venous vessels of rat intestine and mesentery [12,18]. In humans, differentiated responses of sympathetic sub-systems to baroreceptor stimulation have been described which are similar to those reported here: microneurographic studies revealed substantial decreases in MVC activity particularly during phasic baroreceptor stimuli, whereas the same stimuli had no influence on the activity in CVC neurons [30,31]. Hemodynamic measurements have demonstrated carotid baroreceptor-induced splanchnic vasoconstriction [1] Postganglionic MVC and preganglionic VVC neurons are under strong inhibitory control of the carotid baroreceptors [3,4] and we have shown in the present study that the responses to carotid sinus p,essures of different magnitude do not quantitatively differ between these groups of sympathetic neuron. Postganglionic W C neurons projecting in the hypogastric nerve also showed strong barosensitivity [21]. Kendrick et al. [22] induced vasoconstriction in skeletal muscle and viscera by graded unloading of the carotid baroreceptors in the cat. This vasoconstriction was compared with those induced by electrical stimulation of the preganglionic MVC and postganglionic VVC axons at graded stimulation frequencies. This comparison showed indirectly thai the increase in activity induced by baroreceptor unloading is quantitatively similar in both populations of vasoconslrictor neurons. In a recent stud~,
248
we reported a very similar reflex component of respiratory modulation of the activity in MVC and VVC neurons which depends on respiration related baroreceptor unloading [6]. Thus, preand postganglionic vasoconstrictor neurons to skeletal muscle and to the viscera are very similar regarding their batosensitivity.
Acknowledgements W e t h a n k Mrs. N a n k e B l u h m a n d E i k e T a l l o n e for e x p e r t t e c h n i c a l a s s i s t a n c e . T h i s w o r k was s u p p o r t e d by t h e D e u t s c h e F o r s c h u n g s g e meinschaft.
References 1 Abboud, F.M., Eckberg, D.L., Johannsen, U.J. and Mark, A.L., Carotid and cardiopulmonary baroreceptor control of splanchnic and forearm vascular resistance during venous pooling in man, J. Physiol., 286 (1979) 173-184. 2 Bahr, R., Bartel, B., Blumberg, H. and J~inig, W., Functional characterization of preganglionie neurons projecting in ~he lumbar splanchnic nerves: neurons regulating motility, J. Auton. Nerv. Syst., 15 (1986) 109-130. 3 Bahr, R., Bartel, B., Blumberg, H. and Jfinig, W., Functional characterization of preganglionic neurons projecting in the lumbar splanchnic nerves: vasoconstrictor neurons, J. Auton. Nerv. Syst., 15 (1986) 131-140. 4 Blumberg, H., J~inig, W., Rieckmann, C. and Szulczyk, P., Baroreceptor and chemoreceptor reflexes in postganglionic neurons supplying skeletal muscle and hairy skin, J. Auton. Nerv. Syst., 2 ~,1980) 223-240. 5 Boczek-Funcke, A., H~ibler, H.-J., J~inig,W. and Michaelis, M., Responses to graded baroreceptor stimuli of lumbar vasoconstrictor neurons supplying different vascular beds in the cat, Pfliigers Arch., 419 (1991) R44. 6 Boczek-Funcke, A., H~ibler, H.-J., J~inig,W. and Michaelis, M., Respiratc, ry modulation of the activity in sympathetic neurones supplying muscle, skin and pelvic organs in the cat, J. Physiol. (Lond.), 449 (1992) 333-361. 7 Brender, D. and Webb-Peploe, M.M., Influence of carotid baroreceptors on different components of the vascular system, J. Physiol. (Lond.), 205 (1969) 257-274. 8 Damrmey, R.A.L. and McAllen, R.M., Differential control of sympathetic fibres supplying hindlimb skin and muscle by subretrofacial neurones in the cat, J. Physiol. (Lond.L 395 (1988) 41-56. 9 DiSab,o, J., Parker, P.E., Scott, J.B. and Haddy, F.J., Carotid baroreceptor influence on total and segmental
resistances in skin and muscle vasculatures, Am. J. Physiol., 220 (1971) 1970-1978. 10 Fuxe, K. and Sedvall, G., The distribution of adrenergic nerve fibres to the blood vessels in skeletal muscle, Acta Physiol. Scand., 64 (1965) 75-86. 11 Gregor, M., J~inig, W. and Riedel, W., Response pattern of cutaneous postgangiionic neurons to the hindlimb on spinal cord heating and cooling in the cat, Pfliigers Arch., 363 (1976) 135-140. 12 Haase, E.B. and Shoukas, A.A., Carotid sinus baroreceptor reflex control of venular pressure-diameter relations ir~ rat intestine, Am. J. Physiol., 260 (1991) H752-H758. 13 Hadjiminas, J. and Oberg, B., Effects of carotid barcreceptor reflexes on venous tone in skeletal muscle and intestine of the cat, Acta Physiol. Scand., 72 (1968) 518532. 14 Hagbarth, K.E., Hallin, R.G., Hongell, A., Torebj6~k, H.E. and Wallin, B.G., General characteristics of sympathetic activily in human skin nerves, Acta Physiol. Scand., 84 (1972) 164-176. 15 Hagbarth, K.E. and Vallbo, A.B., Pulse and respiratory grouping of sympathetic impulses in human muscle nerves, Acta Physiol. Scand., 74 (1968) 96-108. 16 Hainsworth, R. and Karim, F., Responses of abdominal vascular capacitance in the anaesthetised dog to changes in carotid sinus pressure, J. Physiol. (Lond.), 262 (1976) 659-677. 17 Hainsworth, R., Karim, F., McGregor, K.H. and Wood, L.M., Hind-limb vascular-capacitance responses in anaesthetized dogs, J Physiol. (Lond.), 337 (1983) 417-428. 18 Hdbert, M.T. and Marshall, J.M., Direct observations of effects of baroreceptor stimulation on mesenteric circulation of the rat, J. Physiol. (Lond~, 400 (1988) 29-44. 19 Janig, W., Organisation of the lumbar sympathetic outflow to skeletal muscle and skin o', the cat hindlimb and tail, Rev. Physiol. Biochem. Pharmacol., 102 (1985) 119-213. 20 Jfinig, W., Pre- and postge,nglionic vasoconstrictor neurons: differentiation, types, and discharge properties, Atanu. Rev. Physiol., 50 (1988) 525-539. 21 J~inig, W., Schmidt, M., Schnitzler, A. and Wesselmann, U., Differentiation of sympathetic neurons projecting in the hypogastrie nerves in terms of their discharge patterns ix cats, J. Physiol., 437 (1991) 157-179. 22 Kendrik, E., Oberg, B. and Wennergren, G., Vasoconstrictor fibre discharge of skeletal muscle, kidney, intestine and skin at various levels of arterial baroreceptor activity in the cat, Acta Physiol. Scand., 85 (1972) 464-476. 23 Lovick, T.A., Differential control of cardiac and vasomotor activity by neurones in nucleus paragigantocellularis lateralis in the cat, J. Physiol. (Lond.), 389 (1987) 23-35. 24 Marshall, J., The influence of the sympathetic nervous system on individual vessels of the microcirculation of skeletal muscle of the rat, J. Physiol. (Lond.), 332 (1982) 169-182. 25 McAllen, R.M. and Dampney, R.A.L., Vasomotor neurons in the rostral ventrolateral medulla are organized
~9 topographically with respect to type of vascular bed and not body region, Neurosci. Lett., I l0 (1990) 91-96. 26 Morrison, S.F. and Reis, D.J., Responses of sympathetic preganglionic neurons to rostral ventrolateral medullary stimulation, Am. J. Physiol., 261 (1991) R1247-R1256. 27 Ninomiya, J., Nisimaru, N. and lrisawa, H., Sympathetic nerve activity to spleen, kidney and heart in response to baroreceptor input, Am, J. Physiol., 221 (1971) 1346-1351. 28 Ninomiya, J. and Irisawa, H., Noa-uniformity of symw~thetic nerve activity in response to baroreceptor inpt~ts, Brain Res., 87 (1975) 313-322. 29 Sagawa, K. and Watanabe, K., Summation of bila':eral carotid sinus signals in the barostatic reflex, Am. J. Physiol., 209 (1965) 1278-1286.
30 Wallin, B.G. and Eckberg, D.L., Sympathetic trans!ents caused by abrupt alterations of carotid baroreceptor activity in humans, Am. J. Physiol., 242 (1982) H!85-tt190. 31 Wallin, B.G., Sundl6f, G and Delius, W.. The effect of carotid sinus nerve stimulation on muscle and skin nerve sympathetic activity, in man, Pfliigers Arch., 358 (1975) 101-110. 32 Webb-PepWoe,M.M. and Shepherd, J.T., Response of large hindlimb veins of the dog to sympathetic nerve stimulation, Am. J. Physiol., 215 (1968) 299-?,07. 33 Wennergren, G., Aspects of central integrative and efferent mechanisms in cardiovascular reflex control, Acta Physiol. Scand., Suppl. 428 (1975) 1-53,
241
JANS 01369
Responses of lumbar vasoconstrictor neurons supplying different vascular beds to graded baroreceptor stimuli in the cat M. M i c h a e l i s , A . B o c z e k - F u n c k e , H . - J . H h b l e r a n d W . J[inig Physiologisches Ir..stitut, Christian-Albrechts-Universitiit, Kiel, FRG
(Received 4 August 1992) (Reviston received 14 October 1992) (Accepted 14 October 1992)
K ~ , words': Sympathetic vasoconstrictor neurons; B a r o r e c e p t o r reflex; Cat
Abstract Lumbar ~ympathetic vasoconstrictor neurons supplying skeletal muscle, hai~ skLn. and pelvic organs were tested for their responses to carotid haroreceptor stimulation in ehloralose-anaesthetized cats. Us:ng single- and few-fibre recordings, the responses of the different types of vasoconstrictor neuron to graded steps of non-pulsati;e pressure ranging from 110 to 260 mmHg in a vascularly isolated carotid sinus were analyzed quantitatively during the first 10 s of stimulation. The activity in all postganglionic muscle vasoconstrictor (MVC) neurons, preganglionic visceral vasoconstrictor (VVC) neurons and one third of the poszganglionic cutaneous vasoconstrictor (CVC 1) neurons was strongly depressed by maximal baroreceptor stimulation. Moreover, quantitative analysis revealed no significant differences of the baroreceptor sensitivity of MVC and CVC,. neurons as compared with VVC neurons at all levels of carotid sinus pressure. In contrast, two-thirds of the postganglionic cutaneous vasoconstrictor (CVC 2) neurons exhibited a significantly weaker barosensitivity. The functional implications are discussed.
Introduction T h e s p o n t a n e o u s activity in many pre- a n d postganglionic sympathetic n e u r o n s is u n d e r inhibitory control of arterial baroreceptors. However, d e t e c t a b l e barosensitivity is not a c o m m o n f e a t u r e o f all sympathetic neurons. Thus, the activity in only about 25% o f the preganglionic sympathetic ne'.:,ro~s projecting into the l u m b a r splanchnic nerves is inhibited by b a r o r e c e p t o r stimulation during the systolic rise of the pulse p r e s s u r e wave or large pressure increases in an isolated carotid blind sac [2,3]. On the o t h e r
Correspondence to: M. Michaelis, Physiologisches Institut, Christian-AIbrechts-Universit~it, OIshausenstr. 40, D-2300 Kiel, FRG.
hand, all postganglionic sympathetic neurons with s p o n t a n e o u s activity projecting to skeletal muscle of the hindlimb show strong barosensitivity. These neurons very likely form a functionally homogeneous group of muscle vasoconstrictor (MVC) neurons [4]. Although the postganglionic sympathetic fibres projecting to hairy skin of the hindlimb, which are spontaneously active, also have vasoconstrictor function, only roughly onethird of these neurons exhibits a clearly detectablc barosensitivity [4,19,20]. Similar differences b e t w e e n M V C and cutaneous vasoconstrictor (CVC) n e u r o n s regarding their barosensitivity exist in h u m a n muscle and skin nerves [14,15,31]. N u m e r o u s studies revealed quantitatively diff e r e n t responses of specific vascular beds to baror e c e p t o r stimulation [7,9,13,16,17,22,33]. However, these studies give only indirect information
242 about the neuronal components of these reflex responses. Comparing whole nerve recordings, Ninomiya and coworkers [27,28] described non-uniform responses to baroreceptor stimulation in sympathetic fibres to heart, kidney, spleen and intestine. On one hand this could be due to quantitative differences in the barosensitivity of a uniform population of vasoconstrictor neurons projecting to the various organs. On the other, the recorded nerves could contain different proportions of vasoconstrictor neurons of quantitatively similar barosensitivity. In the present experiments we recorded activity from single- or few-fibre preparations of MVC, CVC and visceral vasoconstrictor neurons and quantitatively studied the responses of these neurons to graded stimulation of the arterial baroreceptors. Some of the results have been published in preliminary form [5].
Materials and Methods
Anaesthesia and animal maintenance Experiments were performed on 12 cats of either sex weighing 2.5-4.7 kg. Anaesthesia was induced by injection of ketamine (Ketanest R, Parke-Davis, 15-20 m g / k g i.m.) and continued with a-o-glucochloralose (a-Chloralose R, Serva, 40 m g / k g i.p.). Supplementary doses of a-D-glucochioralose were given intravenously (10 m g / k g ) to maintain deep anaesthesia as judged from miotic pupils and absence of spontaneous blood pressure and heart rate fluctuations. Systemic arterial blood pressure was continuously recorded using a pressure transducer (List LM-22) connected to a cannula in the femoral or brachial artery. Throughout the experiments mean arterial blood pressure exceeded 90 mmHg. The jugular vein was cannulated for drug administration. Animals were immobilized with pancuronium bromide (Pancuronium-Organon R, Organon Teknika, 0.8 m g / h i.v.) and artificially ventilated through a tracheal cannula keeping the end-expiratory CO z at 3.5-4.5 vol%. Body core temperature was recorded intra-oesophageally and
maintained at 37-38°C by a heating plate. The urinary bladder was catheterized transurethrally to monitor urine excretion. At the end of the experiments the animals were killed by int~avenous injection of a saturated potassium chloride solution under deep anaesthesia. All experiments had been approved by the local animal care committee of the state administration and were conducted in accordance with the German federal law.
Surgery The region of the left carotid sinus was vascularly isolated by ligating the internal carotid artery and the large branches of the external carotid artery. A cannula inserted into the left external carotid artery was connected to a pressure reservoir filled with saline at 25°C and to a pressure transducer (List LM-22) in order to measure the carotid sinus pressure continuously° In all experiments the right carotid sinus nerve was cut. The aortic nerves were left intact. In six experiments the !Left sympathetic trunk and lumbar spianchnic nerves were exposed using a lateral approach, One lumbar splanchnic nerve was dissected free from surrounding tissue and mounted on a perspex platform. Subsequently, exposed tissue and nerves were covered with warm paraffin oil in a pool made out of skin flaps. In a second set of six experiments the deep and superficial peroneal nerves of the left hindlimb were exposed, mounted on perspex platforms and covered with pa~'affin oil. Nerve strands were isolated fiom the lumbar splanchnic nerves, cut peripherally and split until single unit activity could be recorded. Strands containing several postganglionic axons with spontaneous activity were dissected from those parts of the deep peroneal nerve which innervate the anterior tibial muscle and the extensor digitorum longus muscle and from the main part of the superficial peroneal nerve which innervates hairy skin.
Recording Neuronal activity was recorded unipolarly with respect to a reference electrode which was attached to the nearby tissue and was amplified by
243
a low-noise AC preamplifier (input impedance 10 MI~) and a main amplifier with low input impedance, filtered with a band width of 80-120 Hz to 1000-1200 Hz.
before the onset of the stimulus (M) and mean arterial blood pressure 18 s after the onset (N):
Baroreceptor stimulation
For statistical comparison of mean decreases in the activity of different types of vasoconstrictor neurons Student's t-test was applied. Average responses of vasoconstrictor neurons were tested for significance with the paired t-test.
The carotid blind sac could be closed reversibly by a snare which was set around the commc, n carotid artery. As a result the carotid sinus pressure in the closed blind sac fell to 3 0 - ! 0 0 mmHg whilst the systemic blood pressure markedly increased (see Fig. 2B). This residual non-pulsatile pressure was presumably due to retrograde blood flow through small untied arteries. The carotid baroreceptors were stimulated by stepwise raising the non-pulsatile pressure in the closed blind sac to 110-260 mmHg keeping the pressure level constant for at least 10 s. Between two stimuli the carotid sinus pressure was lowered to the residual level. The stimulation protocol usually consisted of a series of five different pressure steps with differences of 30 mmHg, starting with a stimulus in the range of 110-140 mmHg up to a stimulus in the range of 230-260 mmHg.
Data analysis Neuronal activity, carotid sinus pressure, mean arterial blood pressure and ECG were stored on magnetic tape (EMI SE 7000). During .off-line analysis, neuronal activity was fed through window discriminators to produce standard impulses. The decrease in neuronal activity and the reduction of mean arterial blood pressure induced by stimulation of carotid baroreceptors were quantified using a computer program. The relative decrease in activity ( R D A ) was calculated from the relation between the summed neuronal activity during 10 s before the onset of the stimulus (A) and the summed neuronal activity of the first 10 s after onset of the stimulus (B): R D A = ( A - B ) A - t 100%. The relative decrease in blood pressure ( R D B ) was calculated from the relation between the mean arterial blood pressure during the 10 s
RDB=(M-N)M-IIO0%.
Results
Functional identification of vasoconstrictor neurOllS
We recorded activity from eight filaments containing few (3-7) postganglionic units with spontaneous activity projecting to skeletal muscle of the hindlimb. In all cases the activity showed a strong ECG-related rhythmic modulation (cardiac rhythmicity). These neurons will be referred to as muscle vasoconstrictor (MVC) neurons. Activity in postganglionic sympathetic neurons projecting to hairy skin of the hindlimb was recorded from thirteen nerve strands (nine fewfibre preparations, four single fibres). These neurons were tentatively classified as cutaneous vasoconstrictor (CVC) neurons for two reasons: vasodilator neurons to hairy skin exhibit no spontaneous activity and pilomotor and sudomotor neurons innervating the hairy skin of the hindlimb most likely do not exist (see [19]). The activity in four out of 13 CVC preparations showed a strong cardiac rhythmicity, whereas no or weak pulse synchronous modulation could be detected in the activity of the remaining CVC neurons. This proportion is consistent with the proportion of CVC neurons showing strong cardiac rhythmicity reported in a previous study [4]. Preganglionic neurons projecting in the lumbar splanchnic nerves were classified as visceral vasoconstrictor (VVC) neurons when they were under clear inhibitory control of the arterial baroreceptors (see [3]). In the present study the activity in seven VVC neurons was analysed. Figure IA shows a typical example. The activity in this VVC neuron exhibited a strong cardiac rhythmicity and
244
was d e p r e s s e d during a high pressure stimulus in the carotid sinus blind sac (Fig. 1A). T h e majority of preganglionic n e u r o n s projecting in the lumbar splanchnic nerves did not res p o n d to stimulation o f the arterial b a r o r e c e p t o r s (Fig. 1B). T h e s e n e u r o n s are p r o b a b l y involved in regulation of colon and pelvic o r g a n motility a n d are t h e r e f o r e called motility-regulating ( M R ) n e u r o n s [2].
Quantitatiz'e analysis
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2rain Fig. 2 Responses of a single preganglionie visceral vasoconstrictor neuron (VVC) to graded isotonic; pressure stimuli applied to the carotid baroreceptors. (A): response to maximal baroreceptor stimulus. Traces from above: neuronal activity, carotid sinus pressure (CSP) and blood pressure (BP). Regular blood pressure oscillations are induced by artificial ventik:tion. (B): neuronal activity given as a peristimulus-histogram, bin width 2 s (VVC). After closure of the carotid blind sac (downward arrow) CSP dropped simultaneously with the excitation of the unit while the mean arterial blood pressure (MAP) rose. Opening of the carotid blind sac (upward arrow) led to a brief marked inhibition of the unit and to a decline of MAP to resting level, Note the graded inhibition of neuronal activity depending on the magnitude of the baroreceptor stimulus. The pressure stimulus marked with an asterisk is shown as original record in A. (C): post-R-wave histogram (bin width 8 ms), 500 cycles superimposed.
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Fig. 1. Responses of a single preganglionic visceral vasoconstrictor neuron (A), a single preganglionic motility regulating neuron (B) and blood pressure to isotonic pressure stimuli applied to the carotid sinus baroreceptors. Traces from above show carotid sinus pressure (CSP), mean arterial blood pressure (MAP) and neuronal activity in the visceral vasoconstrictor neuron ( W C ) and the motility regulating neuron (MR; peristimulus-histograms, bin width 2 s). Insets show the discriminated actio~ potentials of the units, five times superimposed. The post-R-wave histograms (bin width 8 ms) were obtained from 500 superimposed sweeps over a period of two cardiac cycles.
plied to t h e carotid sinus blind sac w e r e d e t e r mined. As o u t l i n e d in M a t e r i a l s a n d M e t h o d s the residual i n t r a c a r o t i d p r e s s u r e r a n g e d b e t w e e n 30 and 100 m m H g . T h e m e a n residual values did not differ b e t w e e n the two e x p e r i m e n t a l series: 50.3 + 15.9 m m H g during M V C / C V C recordings a n d 60.0 + 11.9 m m H g during V V C recordings ( P > 0.2). A typical e x p e r i m e n t is shown in Fig. 2. Each r a p i d rise of carotid sinus p r e s s u r e was followed by a p r o m p t d e c r e a s e in the activity o f
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Fig. 3. Quantified decreases in the activity of muscle (MVC), visceral (VVC) and cutaneous (CVCI, CVC 2) vasoconstrictor neurons to graded pressure steps applied to the isolated carotid sinus. Dots represent the individual data. All MVC, VVC. CVC z ~nd four of nine CVC 2 preparations were tested with five different baroreceplor stimuli each. Five CVC 2 were only tested with raaximal baroreceptor stimuli > 230 mmltg. CSP, carotid sinus pressure.
the single VVC neuron and by a concomitant fall of blood pressure (Fig. 2B). Both were dependent on the magnitude of the stimulus. Neuronal responses and induced blood pressure changes were quantified (see Materials and Methods). Maximal baroreceptor stimuli (carotid sinus pressure above 230 mmHg) depressed almost totally the activity, in all MVC and VVC neurons (Table 1), whereas CVC neurons did not respond homogeneously to this stimulus. The activity in four out of 13 CVC preparations decreased by 66% or more (CVCt; Table I). These CVC~ neurons also showed a strong cardiac rhythmicity in their a&ivity. Compared with the responses of the remaining CVC neurons (CVCz), the decreases in the activity ia CVC~ neurons as well as those in MVC and VVC neurons were significantly stronger (t-test, P < 0.001; Table I).
MVC, VVC and CVC t neurons 6xhibited no significant differences in their responses to maximal baroreceptor stimulation. Figure 3 illustrates the quantitative responses separately for the different types of vasoconstricTABLE I Ra . . . . . . . . ~f ,4ifferp,,t h ..... o f . . . . . . . . . . ;. . . . neurons to maxim a l stimulation o f carotid baroreceptors
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tor neuron. All MVC, and CVC t and four out o f nine C V C 2 p r e p a r a t i o n s were tested with five graded isotonic pressure stimuli. Five CVC preparations only tested with maximal b a r o r e c e p tor Stimuli decreased in their activity by less than 50% and were t h e r e f o r e included in the g r o u p of C V C 2 neurons. T h e responses of M V C neurons to carotid sinus pressure stimuli exceeding 140 m m H g w e r e significant relative to control activity (paired ttest, P < 0 . 0 5 ) . Carotid pressures of 110-140 m m H g induced no significant responses. Like M V C neurons, W C and CVC~ neurons exhibited significant decreases in their activity to
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carotid sinus pressures o f 140-170 m m H g and more. In contrast, CVC 2 n e u r o n s showed n o significant responses except to maximal b a r o r e c e p t o r stimuli (230-260 m m H g ) . Figure 4 depicts the a v e r a g e d data. F o r the different types o f vasoconstrictor n e u r o n the m e a n decreases in activity were calculated separately for five 3 0 - m m H g carotid sinus pressure intervals beginning with the 110-140 m m H g interval. T h e d e c r e a s e s in m e a n arterial b l o o d pressure w e r e calculated in the same way. Statistical analysis revealed no significant d i f f e r e n c e s b e t w e e n the responses in MVC, V V C a n d CVC~ neurons. However, the b a r o r e c e p t o r - i n d u c e d decreases in these t h r e e types of vasoconstrictor n e u r o n w e r e significantly g r e a t e r than those in C V C 2 n e u r o n s regarding carotid sinus pressures exceeding 170 m m H g (t-test, P < 0.05). T h e d e c r e a s e s in m e a n arterial b l o o d pressure i n d u c e d by the same stimuli in e x p e r i m e n t s in which n e u r o n a l activity was r e c o r d e d f r o m V V C alone, and simultaneously from M V C and CVC 1, and f r o m M V C and C V C 2, respectively, did not differ significantly.
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Fig. 4. Mean decreases in neuronal activity, obtained from different types of vasoconstrictor neuron, and concomitant mean decreases in mean arterial blood pressure (MAP) induced by graded isotonic baroreceptor stimuli (CSP, carotid sinus pressure). Individual data were averaged from intervals of 3U mmi-ig each. Vertical bars give 1 S.E.M. Mean responses in MVC neurons (®), neurons (#) and CVC l neurons ( I ) d i d not differ significantly from each other (t-test, P > 0.05). The mean decreases in activity in these three types of vasoconstrictor neurons were significantly greater than those in CVCe neurons (D) regarding carotid sinus pressures in excess of 170 mmHg (t-test, P < 0.05). Activity in MVC neurons was always recorded simultaneously with activity in CVC~ or CVCz neurons. Therefore, only the mean decreases in MAP obtained during recording from CVC~ (m) ana CVC 2 (12) neurons were given. Mean decreases in MAP during recording from CVC~, CVC 2 and VVC neurons ( q ) were not significantlydifferent.
WC
In the p r e s e n t study we described the quantitative responses of different types o f vasoconstrictor n e u r o n to b a r o r e c e p t o r stimulation using an isolated carotid blind sac p r e p a r a t i o n . Most likely the o b s e r v e d changes in sympathetic activity during increased carotid sinus pressures were i n d e e d due to b a r o r e c e p t o r stimulation and not due to a c o n c o m i t a n t c h e m o r e c e p t o r activation. It is known that the activity in C V C n e u r o n s without cardiac rhythmicity is inhibited by c h e m o r e c e p t o r stimulation [4]. In our study the C V C 2 n e u r o n s which s h o w e d no cardiac rhythmicity were not or only weakly inhibited by p r e s s u r e steps in the carotid blind sac. Thus, if these pressure steps would lead to a significant c h e m o r e c e p t o r activation, o n e would expect a m o r e p r o m i n e n t inhibitioJa in C V C 2 neurons. F u r t h e r m o r e , M V C neurons are strongly activated by c h e m o r e c e p t o r stimulation [4] but we o b s e r v e d an almost complete d e p r e s s i o n in their activity during large distension o f the isolated carotid sinus.
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Significant responses of MVC, VVC and C V C l neurons were detected using carotid sinus pressures of more than 140 mmHg. This relatively high intensity may have several reasons. First, compared with synchronous stimulation of carotid baroreceptor afferents on both sides, pressure steps in an unilateral carotid blind sac induce only half as large decreases in systemic blood pressure [29]. The same applies probably to the induced decreases in vasoconstrictor activity. Secondly, we left the aortic nerves intact. Thus, unloading of aortic baroreceptor affe rents during the decrease of systolic blood pressure may have partly counteracted the reflex inhibition of vasoconstrictor neurons during carotid sinus pressure stimuli. There is strong evidence that bulbospinal neurons located in the rostral ventrolateral medulla are important for the control of the activity in vasoconstrictor neurons. Furthermore, there seem to be several subpopulations of 'presympathetic' neurons which may be associated with the regulation of different vascular beds [8,23,25,26]. Thus, the differences and similarities of baroreceptor sensitivity found in different types of sympathetic vasoconstrictor neurons may reflect the specific medullary organization of presympathetic neurons. It is known from anatc,mical investigations that veins in the skeletal ml~scle, in contrast to arteries, receive no significant sympathetic innervation [10,24]. Thus, MVC neurons presumably innervate almost exclusively resistance vessels. MVC neurons are under strong inhibitory control of arterial barorecaptors as shown in this and other investigations [4,20]. It is likely that strong baroreceptor sensitivity is not only typical for MVC neurons but is a common feature of postganglionic sympathetic neurons innervating resistance vessels. Thus, CVC~ neurons which showed the same barosensitivity as MVC neurons may innervate the resistance vessels in the hairy skin. This would be consistent with results of DiSalvo et al. [9]. Measuring "the changes of resistance they observed no differences between dilatation of small muscle and skin arteries during baroreceptor stimulation. Vasoconstriction of peripheral superficial but not of proximal and deep
veins of the hindlimb could be observed during electrical stimulation of the lumbar sympathetic trunk [17,32]. However, baroreceptor stimulatior~ does not induce a significant diameter change of peripheral cutaneous veins [7,17] indicating that cutaneous vasoconstrictor neurons supplying these veins are not under significant baroreceptor control. Therefore, CVC 2 neurons may innervate peripheral cutaneous veins. Furthermore, CVC 2 neurons may play a role in thermoregulatien [i i]. VVC neurons may project to resistance as well as capacitance vessels in the visceral domain. Stimulation of arterial baroreceptors leads to a clear reflex change in abdominal vascular resistance and capacitance [16,22]. Direct observations of blood vessels during baroreceptor stiraulation have revealed significant diameter-changes in both arterial and venous vessels of rat intestine and mesentery [12,18]. In humans, differentiated responses of sympathetic sub-systems to baroreceptor stimulation have been described which are similar to those reported here: microneurographic studies revealed substantial decreases in MVC activity particularly during phasic baroreceptor stimuli, whereas the same stimuli had no influence on the activity in CVC neurons [30,31]. Hemodynamic measurements have demonstrated carotid baroreceptor-induced splanchnic vasoconstriction [1] Postganglionic MVC and preganglionic VVC neurons are under strong inhibitory control of the carotid baroreceptors [3,4] and we have shown in the present study that the responses to carotid sinus p,essures of different magnitude do not quantitatively differ between these groups of sympathetic neuron. Postganglionic W C neurons projecting in the hypogastric nerve also showed strong barosensitivity [21]. Kendrick et al. [22] induced vasoconstriction in skeletal muscle and viscera by graded unloading of the carotid baroreceptors in the cat. This vasoconstriction was compared with those induced by electrical stimulation of the preganglionic MVC and postganglionic VVC axons at graded stimulation frequencies. This comparison showed indirectly thai the increase in activity induced by baroreceptor unloading is quantitatively similar in both populations of vasoconslrictor neurons. In a recent stud~,
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we reported a very similar reflex component of respiratory modulation of the activity in MVC and VVC neurons which depends on respiration related baroreceptor unloading [6]. Thus, preand postganglionic vasoconstrictor neurons to skeletal muscle and to the viscera are very similar regarding their batosensitivity.
Acknowledgements W e t h a n k Mrs. N a n k e B l u h m a n d E i k e T a l l o n e for e x p e r t t e c h n i c a l a s s i s t a n c e . T h i s w o r k was s u p p o r t e d by t h e D e u t s c h e F o r s c h u n g s g e meinschaft.
References 1 Abboud, F.M., Eckberg, D.L., Johannsen, U.J. and Mark, A.L., Carotid and cardiopulmonary baroreceptor control of splanchnic and forearm vascular resistance during venous pooling in man, J. Physiol., 286 (1979) 173-184. 2 Bahr, R., Bartel, B., Blumberg, H. and J~inig, W., Functional characterization of preganglionie neurons projecting in ~he lumbar splanchnic nerves: neurons regulating motility, J. Auton. Nerv. Syst., 15 (1986) 109-130. 3 Bahr, R., Bartel, B., Blumberg, H. and Jfinig, W., Functional characterization of preganglionic neurons projecting in the lumbar splanchnic nerves: vasoconstrictor neurons, J. Auton. Nerv. Syst., 15 (1986) 131-140. 4 Blumberg, H., J~inig, W., Rieckmann, C. and Szulczyk, P., Baroreceptor and chemoreceptor reflexes in postganglionic neurons supplying skeletal muscle and hairy skin, J. Auton. Nerv. Syst., 2 ~,1980) 223-240. 5 Boczek-Funcke, A., H~ibler, H.-J., J~inig,W. and Michaelis, M., Responses to graded baroreceptor stimuli of lumbar vasoconstrictor neurons supplying different vascular beds in the cat, Pfliigers Arch., 419 (1991) R44. 6 Boczek-Funcke, A., H~ibler, H.-J., J~inig,W. and Michaelis, M., Respiratc, ry modulation of the activity in sympathetic neurones supplying muscle, skin and pelvic organs in the cat, J. Physiol. (Lond.), 449 (1992) 333-361. 7 Brender, D. and Webb-Peploe, M.M., Influence of carotid baroreceptors on different components of the vascular system, J. Physiol. (Lond.), 205 (1969) 257-274. 8 Damrmey, R.A.L. and McAllen, R.M., Differential control of sympathetic fibres supplying hindlimb skin and muscle by subretrofacial neurones in the cat, J. Physiol. (Lond.L 395 (1988) 41-56. 9 DiSab,o, J., Parker, P.E., Scott, J.B. and Haddy, F.J., Carotid baroreceptor influence on total and segmental
resistances in skin and muscle vasculatures, Am. J. Physiol., 220 (1971) 1970-1978. 10 Fuxe, K. and Sedvall, G., The distribution of adrenergic nerve fibres to the blood vessels in skeletal muscle, Acta Physiol. Scand., 64 (1965) 75-86. 11 Gregor, M., J~inig, W. and Riedel, W., Response pattern of cutaneous postgangiionic neurons to the hindlimb on spinal cord heating and cooling in the cat, Pfliigers Arch., 363 (1976) 135-140. 12 Haase, E.B. and Shoukas, A.A., Carotid sinus baroreceptor reflex control of venular pressure-diameter relations ir~ rat intestine, Am. J. Physiol., 260 (1991) H752-H758. 13 Hadjiminas, J. and Oberg, B., Effects of carotid barcreceptor reflexes on venous tone in skeletal muscle and intestine of the cat, Acta Physiol. Scand., 72 (1968) 518532. 14 Hagbarth, K.E., Hallin, R.G., Hongell, A., Torebj6~k, H.E. and Wallin, B.G., General characteristics of sympathetic activily in human skin nerves, Acta Physiol. Scand., 84 (1972) 164-176. 15 Hagbarth, K.E. and Vallbo, A.B., Pulse and respiratory grouping of sympathetic impulses in human muscle nerves, Acta Physiol. Scand., 74 (1968) 96-108. 16 Hainsworth, R. and Karim, F., Responses of abdominal vascular capacitance in the anaesthetised dog to changes in carotid sinus pressure, J. Physiol. (Lond.), 262 (1976) 659-677. 17 Hainsworth, R., Karim, F., McGregor, K.H. and Wood, L.M., Hind-limb vascular-capacitance responses in anaesthetized dogs, J Physiol. (Lond.), 337 (1983) 417-428. 18 Hdbert, M.T. and Marshall, J.M., Direct observations of effects of baroreceptor stimulation on mesenteric circulation of the rat, J. Physiol. (Lond~, 400 (1988) 29-44. 19 Janig, W., Organisation of the lumbar sympathetic outflow to skeletal muscle and skin o', the cat hindlimb and tail, Rev. Physiol. Biochem. Pharmacol., 102 (1985) 119-213. 20 Jfinig, W., Pre- and postge,nglionic vasoconstrictor neurons: differentiation, types, and discharge properties, Atanu. Rev. Physiol., 50 (1988) 525-539. 21 J~inig, W., Schmidt, M., Schnitzler, A. and Wesselmann, U., Differentiation of sympathetic neurons projecting in the hypogastrie nerves in terms of their discharge patterns ix cats, J. Physiol., 437 (1991) 157-179. 22 Kendrik, E., Oberg, B. and Wennergren, G., Vasoconstrictor fibre discharge of skeletal muscle, kidney, intestine and skin at various levels of arterial baroreceptor activity in the cat, Acta Physiol. Scand., 85 (1972) 464-476. 23 Lovick, T.A., Differential control of cardiac and vasomotor activity by neurones in nucleus paragigantocellularis lateralis in the cat, J. Physiol. (Lond.), 389 (1987) 23-35. 24 Marshall, J., The influence of the sympathetic nervous system on individual vessels of the microcirculation of skeletal muscle of the rat, J. Physiol. (Lond.), 332 (1982) 169-182. 25 McAllen, R.M. and Dampney, R.A.L., Vasomotor neurons in the rostral ventrolateral medulla are organized
~9 topographically with respect to type of vascular bed and not body region, Neurosci. Lett., I l0 (1990) 91-96. 26 Morrison, S.F. and Reis, D.J., Responses of sympathetic preganglionic neurons to rostral ventrolateral medullary stimulation, Am. J. Physiol., 261 (1991) R1247-R1256. 27 Ninomiya, J., Nisimaru, N. and lrisawa, H., Sympathetic nerve activity to spleen, kidney and heart in response to baroreceptor input, Am, J. Physiol., 221 (1971) 1346-1351. 28 Ninomiya, J. and Irisawa, H., Noa-uniformity of symw~thetic nerve activity in response to baroreceptor inpt~ts, Brain Res., 87 (1975) 313-322. 29 Sagawa, K. and Watanabe, K., Summation of bila':eral carotid sinus signals in the barostatic reflex, Am. J. Physiol., 209 (1965) 1278-1286.
30 Wallin, B.G. and Eckberg, D.L., Sympathetic trans!ents caused by abrupt alterations of carotid baroreceptor activity in humans, Am. J. Physiol., 242 (1982) H!85-tt190. 31 Wallin, B.G., Sundl6f, G and Delius, W.. The effect of carotid sinus nerve stimulation on muscle and skin nerve sympathetic activity, in man, Pfliigers Arch., 358 (1975) 101-110. 32 Webb-PepWoe,M.M. and Shepherd, J.T., Response of large hindlimb veins of the dog to sympathetic nerve stimulation, Am. J. Physiol., 215 (1968) 299-?,07. 33 Wennergren, G., Aspects of central integrative and efferent mechanisms in cardiovascular reflex control, Acta Physiol. Scand., Suppl. 428 (1975) 1-53,