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In vagotomized, midcollicular decerebrate, non-anesthetized cats, in which the preganglionic input to the stellate ganglion was intact and the right forelimb was autoperfused at constant flow, the muscarinic receptor antagonist pirenzepine (PZP, 50 micrograms/kg i.v.), which does not cross the blood-brain barrier, produced a decrease in forelimb perfusion pressure (FLPP), heart rate (HR) and systemic arterial pressure (SAP). Administration of the nicotinic receptor antagonist hexamethonium (30 mg/kg i.v.) after PZP produced a further, larger drop in FLPP, HR and SAP. The dose of PZP used blocked the increase in FLPP and HR evoked by the muscarinic receptor agonist McN-A-343, but not the increase evoked by the nicotinic receptor agonist DMPP, when these drugs were injected into the arterial supply of the ganglion. Following administration of phentolamine and propranolol (2 mg/kg i.v. of each) which caused bradycardia and forelimb vasodilation, PZP had no effect on FLPP and HR. This finding suggests that PZP decreases sympathetic tone in resistance vessels and in the heart. Pirenzepine did not depress the forelimb vasoconstriction and the cardioacceleration evoked by electrical stimulation of the postganglionic vertebral and inferior cardiac nerves, respectively, suggesting that PZP does not act at the neuro-effector junctions. On the other hand, PZP blocked the forelimb vasoconstriction and the cardioacceleration produced by stimulating the preganglionic input of the stellate ganglion in presence of hexamethonium (30 mg/kg i.v.), indicating the stellate ganglion as the likely site of action of the drug. These findings suggest that the ganglion cell firing that underlies the sympathetic tone of cardiovascular effector cells is generated, in part, by a muscarinic, PZP-sensitive, synaptic mechanism.

A low-frequency stimulus train to the preganglionic input inhibits synaptic transmission in the superior cervical ganglion (SCG) of the cat. The inhibition is blocked by naloxone as well as by selective antagonists at mu and delta opiate receptors, which suggests that the mediator is an endogenous opioid [27,29]. Exogenous opioid peptides, including methionine-enkephalin (Met-Enk), which is present in preganglionic axons of the SCG, inhibit ganglionic transmission by a naloxone-sensitive mechanism. In the present study we test, in the anesthetized cat, whether the naloxone-sensitive synaptic inhibition is mediated by a pre- and/or post-synaptic mechanism. As a test of presynaptic inhibition, we measured the acetylcholine (ACh) released by preganglionic stimulation into the venous effluent of the perfused SCG. As a test of post-synaptic inhibition, we measured the effect of a preganglionic conditioning train on the ganglion cell firing evoked by ganglion-stimulant drugs injected into the arterial supply of the ganglion. In presence of naloxone (3 microM), which blocked the synaptic inhibition, the amount of ACh released by stimulated preganglionic axons did not change. Thus, the endogenous opioid which mediates the naloxone-sensitive inhibition does not act by depressing ACh release. In contrast, the ganglion cell firing evoked by ganglion-stimulant drugs was markedly depressed by a conditioning train, and naloxone blocked the depression, which suggests that the endogenous mediator of the naloxone-sensitive inhibition acts postsynaptically to decrease the excitability of ganglion cells. Exogenous Met-Enk depressed both ACh release by preganglionic stimulation and the firing of ganglion cells evoked by ganglion-stimulant drugs.(ABSTRACT TRUNCATED AT 250 WORDS)

In anesthetized cats, in which a nodose-superior cervical ganglion (SCG) anastomosis had been performed 6-9 months earlier, the nictitating membrane contraction evoked by electrical stimulation of the cervical vagus nerve ipsilateral to the anastomosis was recorded. The competence of nodose neurons in regulation the multiple synaptic mechanisms of the sympathetic ganglion was tested by comparing this response with the responses to stimulation of (self-reinnervated SCG). The response of the nictitating membrane ipsilateral to the anastomosis was smaller and had a lower EC50 for hexamethonium (C6) than the responses of the nictitating membrane ipsilateral to the intact or sutured CST. A 40 Hz 10s stimulus train to the intact or sutured CST produced potentiation of ganglionic transmission lasting 1 hour or longer, while a similar stimulus train to the anastomosed cervical vagus nerve produced no potentiation. During block of ganglionic nicotinic transmission with C6, CST or vagus nerve stimulation evoked responses which increased in amplitude with increasing stimulus frequency and were blocked by the selective muscarinic receptor antagonist pirenzepine. When the anticholinesterase eserine was added, the responses evoked by preganglionic stimulation decreased in amplitude in the intact SCG, as previously shown [7], and in the self-reinnervated SCG. This effect, which is due to inhibition mediated by muscarinic receptors selectively blocked by AF-DX116, was absent in the anastomosed SCG. During block of ganglionic transmission with C6 and atropine, a 40 Hz stimulus train to the intact or to the sutured CST evoked a slow, small amplitude contraction that was enhanced by naloxone. This response, most likely mediated by peptides [6], was absent in the anastomosed SCG.(ABSTRACT TRUNCATED AT 250 WORDS)

We have previously shown that the neurotensin (NT) store in preganglionic axon terminals of the cat stellate ganglion (SG) is reversibly depleted by prolonged preganglionic stimulation. The present study addresses the questions of whether the preganglionic axon terminals release NT in response to depolarizing stimuli in vitro and whether in vivo NT is released by the tonic firing of the sympathetic preganglionic neurons. Slices of the SG of the anaesthetized cat, maintained in oxygenated Ringer solution, released NT. The efflux increased when the K concentration was increased from 5 to 25 or 45 mM or when veratridine was added to the medium. In Ca-free medium, efflux was suppressed. The effect of veratridine was blocked by tetrodotoxin (TTX). In awake, freely moving cats, in which TTX was applied for 4 days to the preganglionic input of the right SG, the NT content of this ganglion doubled by comparison with the left SG. Since NT accumulates proximal to a ligature on the preganglionic input of the SG, the increased NT content is likely to result from suppression of action potential-dependent release while influx into the terminals persists. This result suggests that the steady state of the NT store in sympathetic preganglionic terminals is the result of a steady influx from the soma balanced by action potential-dependent loss, presumably release.

We have previously shown that prolonged preganglionic stimulation (e.g. 40 Hz 20 min) depletes the neurotensin (NT) store of preganglionic axon terminals in the stellate ganglion (SG) of the cat and that replenishment of the store requires several days. The present study investigates the mechanisms which control turnover of the NT store in sympathetic preganglionic axon terminals. NT content of the SG and of the preganglionic axons which innervate it was determined by radioimmunoassay in the anesthetized cat. This study shows that, during the 24 h after 40-Hz 20-min stimulation of the preganglionic input to the SG, the rate of NT accumulation proximal to a ligature on the stimulated input is three times that observed in the control. The poststimulus increase in NT accumulation rate is prevented by treatment with protein-synthesis inhibitors. When the centripetal propagation of action potentials from the stimulus site on the preganglionic axons is prevented by a tetrodotoxin block applied locally during the stimulation period, the poststimulus increase in NT accumulation rate and the replenishment of the store are both prevented. These data suggest that the level of activity of the neuron regulates NT supply to the axon terminals, presumably by regulating NT synthesis. Thus, in the sympathetic preganglionic neuron, the action potential provides a mechanism for matching peptide synthesis to release.

Nicotinic transmission in the superior cervical ganglion of the cat was studied in vivo and in vitro by recording the postganglionic compound action potential evoked, under partial block with hexamethonium, by supramaximal 0.2 Hz stimulation of the cervical sympathetic trunk. The compound action potential was depressed following a stimulus train (5 Hz, 40 s) applied to the same set of axons. The inhibition was antagonized by naloxone, suggesting mediation by endogenous opioids. Agonists selective for mu-, delta- and kappa-opioid receptors, injected into the arterial supply of the in situ ganglion or added to the perfusion fluid in the in vitro ganglion, also produced a naloxone-sensitive inhibition of the compound action potential. The synaptic inhibition was antagonized by the delta-selective antagonist ICI 174,864 and by the mu-selective antagonist CTAP, but not by the kappa-selective antagonist Nor-BNI. These results suggest that all three main types of opioid receptors are present in the ganglion but only mu- and delta-receptors are involved in the inhibition of ganglionic transmission mediated by the endogenous opioids.

We have shown previously that stimulation of the cat cervical sympathetic trunk for 2 h at 40 Hz Depletes the large Dense-cored vesicle Store in synaptic boutons of the superior cervical ganglion and that post-depletion recovery of the store takes several days. In the present study, we examine the properties of the depletion and recovery mechanisms. Invaginations of the plasmalemma suggestive of the exocytosis of dense cores were seen frequently in boutons from stimulated ganglia. The depletion process is calcium dependent: in ganglia perfused with calcium-free Krebs solution no depletion was produced by 40 Hz preganglionic stimulation. The depletion process is rapid: during continuous stimulation of the cervical sympathetic trunk with 40 Hz, depletion observed by the end of 2 h was similar to depletion by the end of the initial 5 min of stimulation. The depletion process is frequency dependent: when the cervical sympathetic trunk was stimulated with a constant number of stimuli, no depletion occurred at the frequency of 2 or 10 Hz, while the frequencies of 20 and 40 Hz produced depletion, which was greater at 40 Hz. Recovery of the large dense-cored vesicle store during the initial 24 h after 10 min of 40 Hz stimulation was faster, and of approximately the same magnitude, than during the succeeding five days. Recovery of the store after stimulus-evoked depletion was prevented by application of colchicine to the cervical sympathetic trunk, which suggests dependence of recovery on fast axonal transport. Large dense-cored vesicles accumulated in the colchicine-treated segment of cervical sympathetic trunk axons. In conclusion, these observations suggest that the stimulus-evoked depletion of large dense-cored vesicle stores in synaptic boutons of the cat superior cervical ganglion is the result of calcium-dependent exocytosis of the large dense-cored vesicle core and that the post-stimulus recovery is critically dependent on microtubule-mediated axonal transport.

The effect of the muscarinic receptor antagonist AF-DX 116 on the inhibitory action of muscarinic agonists and on responses mediated by nicotinic or muscarinic ganglionic transmission was studied in the superior cervical ganglion of the anesthetized cat. The postganglionic compound action potential evoked by cervical sympathetic trunk stimulation was depressed by methacholine or acetylcholine (ACh) injected into the ganglionic arterial supply. The depression was blocked by AF-DX 116. The compound action potentials evoked by preganglionic stimulus trains were also depressed when the intratrain frequency was 2 Hz or greater. This intratrain depression was, however, insensitive to AF-DX 116. The anticholinesterase drug physostigmine markedly enhanced the intratrain depression of the compound action potential. This effect was reversed by AF-DX 116. During nicotinic receptor block with hexamethonium, preganglionic stimulus trains with intratrain frequencies of 5 Hz or greater produced nictitating membrane contractions that could be blocked by the M1 muscarinic receptor antagonist pirenzepine. The amplitude of the contractions increased with frequency and reached a maximum at 20–40 Hz. AF-DX 116 had no effect on these responses. After administration of physostigmine, the amplitude of the nictitating membrane responses decreased with increasing intratrain frequency. AF-DX 116 reversed this effect. The data suggest that, in the superior cervical ganglion, AF-DX 116 sensitive muscarinic receptors which depress synaptic transmission are activated by exogenous agonists but not by the ACh released by the preganglionic axon terminals unless cholinesterase activity is inhibited. This may result from the fact that these receptors, although accessible to agonists injected into the ganglionic arterial supply, are remote from synaptic release sites and that, under normal conditions, cholinesterase activity prevents sufficient activation of these receptors by the nerve-released ACh to result in modulation of synaptic transmission.Key words: muscarinic inhibition, M2 receptors, sympathetic ganglion modulation.

The superior cervical ganglion of rats was perfused with Ringer solution containing hexamethonium to produce a steady, partial, nicotinic block. The compound action potential (CAP) evoked by supramaximal single shock stimulation of the cervical sympathetic trunk (CST) was recorded from the internal carotid nerve. Bolus injection of the protein kinase C (PKC) activators 4 beta-phorbol-12,13-dibutyrate (PDBu) or 4 beta-phorbol-12,13-diacetate (PDAc) produced a marked, prolonged, dose-dependent potentiation of the CAP amplitude (e.g. 90% decay 2 h). A non-PKC activating phorbol ester (PE), 4 alpha-phorbol-12,13-didecanoate, produced no potentiation. The PE-induced potentiation was antagonized by the PKC inhibitor H-7. In addition, after 1 h exposure to PDBu (3 microM) and recovery from the potentiation (e.g. 2-4 h), a second exposure to PDBu or PDAc produced no potentiation. A 5 s 40 Hz supramaximal train to the CST produced a long lasting potentiation of the CAP (long-term potentiation, LTP) as described previously. However, a similar train did not evoke LTP after perfusion for 1 h with PDBu. The train-evoked LTP was depressed by the PKC inhibitor H-7 at a concentration which antagonized the PE-evoked potentiation. These data suggest that (i) PKC activation potentiates nicotinic transmission, and (ii) a component of the train-evoked LTP is mediated by PKC.

A stimulus train to preganglionic axons produces long-term potentiation (LTP) of population responses of sympathetic ganglion cells evoked by the same or by other converging axons. The present study shows that preganglionic axons emerging from the spinal cord in different thoracic rami, and converging onto a common pool of ganglion cells that innervate a single target, differ in their ability to induce LTP. In anesthetized cats under partial nicotinic block with hexamethonium, the nictitating-membrane (NM) contraction evoked by stimulation of the first (T1) and third (T3) thoracic white rami (WR) was recorded. Each ramus produced a contraction of similar amplitude. In contrast, the homosynaptic potentiation produced by a 40-Hz 10-s train differed markedly. T1WR produced a potentiation of duration comparable to that produced by stimulation of the cervical sympathetic trunk, while T3WR produced either no potentiation or a potentiation of much shorter duration. The NM response evoked by T3WR, however, was potentiated by similar extent and duration as was the response evoked by T1WR when the train was applied to T1WR (heterosynaptic LTP). This suggests that the ganglionic synapses made by T3WR possess the mechanism for expressing LTP. Conversely, T3WR was ineffective at potentiating heterosynaptically the NM response evoked by T1WR. These results suggest that the ability to produce or release the LTP inducer varies markedly among sympathetic preganglionic neurons.Key words: preganglionic neuron heterogeneity, long-term potentiation induction, synaptic transmission, neuropeptides.

The properties of the excitatory postsynaptic potential evoked by focal stimulation and of the responses to excitatory amino acids were examined by intracellular recording from sympathetic preganglionic neurons in upper thoracic spinal cord slices of the adult cat. Single stimuli to the region dorsal to the intermedio-lateral nucleus evoked short-latency, presumably monosynaptic, excitatory postsynaptic potentials. The reversal potential of this response was −2.2 mV and became more negative when external Na + or K + concentration was decreased. The excitatory postsynaptic potential was depressed by the non-selective excitatory amino acid receptor antagonist cis -2,3-piperidine dicarboxylic acid and enhanced by a glutamate uptake inhibitor. Thenon- N -methyl- d -aspartate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione abolished the excitatory postsynaptic potential in 72% of neurons. In the remaining neurons, this antagonist only depressed the potential and unmasked a slower component which was abolished by the N -methyl- d -aspartate receptor antagonist d , l -2-amino-5-phosphonovaleric acid. In the presence of tetrodotoxin all neurons tested were depolarized by glutamate or aspartate, as well as by the selective agonists quisqualate, alpha-amino-3-hydroxy-5-methylisoxazole propionic acid, kainate and N -methyl- d -aspartate. The glutamate-evoked depolarization reversed at a membrane potential of −2.0 mV and at a more negative value when external Na + or K + concentration was decreased. The response to alpha-amino-3-hydroxy-5-methylisoxazole propionic acid was abolished by 6-cyano-7-nitro-quinoxaline-2,3-dione in all neurons tested and that to kainate in only one-third of the cells. In the remainder the response to kainate was only slightly depressed by this antagonist. The responses to glutamate and aspartate were only slightly depressed by the combined action of the various amino acid receptor antagonists used. The responses to N -methyl- d -aspartate were abolished by d , l -2-amino-5-phosphonovaleric acid. The punched-out region of the intermedio-lateral nucleus, maintained in vitro , released glutamate and aspartate in the absence of stimulation. Field stimulation (20 Hz) enhanced release by between 40 and 100%. The increase was prevented by superfusion with calcium-free Krebs. It is concluded that excitatory amino acids, acting on both N -methyl- d -aspartate andnon- N -methyl- d -aspartate receptors, but mainly on the latter, are likely mediators of the monosynaptic excitatory postsynaptic potential evoked in sympathetic preganglionic neurons by the stimulated region. The efflux data suggest that glutamate and aspartate are among the mediators.

Neurotensin (NT) within the stellate ganglion (SG) of the cat is present in axon terminals of extrinsic, presumably preganglionic, neurons and may play a role in ganglionic transmission. NT content of the SG, and of the preganglionic axons which innervate it, was determined by radioimmunoassay in the anesthetized cat under various experimental conditions in order to understand the factors determining the size of the ganglionic NT stores. The immunoreactive NT (iNT) from extracts of SG and its preganglionic inputs (white rami T2 and T3, sympathetic trunk between SG and T4 white ramus) coeluted with synthetic NT(1-13) on RP-HPLC. NT accumulated proximal to ligatures on the preganglionic inputs of the SG. The daily rate of axonal transport of NT, estimated from the accumulation, represents 28.7% of the ganglionic stores of NT. Preganglionic stimulation at 2 Hz for 100 min did not change ganglionic NT content. Preganglionic stimulation at 40 Hz reduced the NT content to 70.4 +/- 1.8% of control in 10 min and to 34.7 +/- 4.2% of control in 20 min. Additional 100 min of 40 Hz stimulation produced no further depletion. The residual iNT, which coeluted with NT1-13, presumably represents a pool of unreleasable NT. Post-depletion recovery was complete in 7 days and showed an initial rapid phase over the first 24 h followed by a slower phase over the remaining 6 days. Pepsin treatment, which has previously been shown to generate iNT from NT precursor in liver and other tissues, provided no evidence of the NT precursor in extracts of SG and its preganglionic input.(ABSTRACT TRUNCATED AT 250 WORDS)

In anesthetized, artificially ventilated cats, pretreated with the muscarinic antahonist scopolamine, the effect of naloxone on the efficacy of nicotinic transmission in the superior cervical ganglion was used as a test of endogenous opioid release by the preganglionic axons. The cervical sympathetic trunk (CST) was split into two bundles. The compound action potential (CAP), evoked by supramaximal low-frequency stimulation (0.25 Hz) of one CST bundle, was recorded from a postganglionic nerve of the superior cervical ganglion. Partial block with hexamethonium was used to reduce the ‘safety factor’ of nicotinic transmission. A conditioning train (5 Hz, 40 s, supramaximal) to the other CST bundle (heterosynaptic conditioning) inhibited the CAP. At the peak of the inhibition the CAP was attenuated by 51 ± 4%. Recovery was 90% complete in 201 ± 28s. The inhibition was antagonized in a dose-dependent manner by i.v. naloxone with an apparent IC 50 of 60 ± 12 μg/kg. The maximum effect obtained with naloxone was an 80% decrease in the magnitude of the inhibition. Magnitude and duration of the heterosynaptic inhibition were related to frequency, duration and intensity of the conditioning train. Naloxone-sensitive inhibition was observed with frequencies of the conditioning train as low as 0.5 Hz. A train (5 Hz, 40 s) to the postganglionic nerves produced a naloxone-insensitive depression of the orthodromic test CAP which was of smaller amplitude and duration than when the conditioning train was applied to the preganglionic axons. The endopeptidase inhibitors thiorphan (0.3–0.5 mg/kg i.v.) and captopril (0.2 mg/kg i.v.) increased magnitude and duration of the depression produced by CST stimulation, but not of that produced by postganglionic nerve stimulation. The aminopeptidase inhibitor bestatin (0.2 mg/kg i.v.) had no effect. These observations suggest that preganglionic axon terminals in the superior cervical ganglion release opioid peptides when stimulated electrically. Very low frequencies of preganglionic stimulation are effective, suggesting that endogenous opioids may also be released during the natural discharge of the preganglionic neurons. Endopeptidases provide a significant modulation of the actions of the endogenous opioids.

1. In the anesthetized, scopolamine-treated cat, the compound action potential (CAP) evoked by a single supramaximal shock to the third thoracic white ramus (T3WR) was recorded in the inferior cardiac nerve (ICN). The CAP was depressed in a dose-dependent manner by the intravenous administration of the nicotinic antagonist hexamethonium (C6). 2. During steady intravenous infusion of C6, which reduced the amplitude of the CAP by 80-90%, a short train of stimuli (few seconds, 10-40 Hz) to the sympathetic trunk just below T4WR potentiated the CAP for periods of tens of minutes to 1-2 h (heterosynaptic long-term potentiation, LTP). An LTP of similar time course was obtained when both train and single shock were applied to T3WR (homosynaptic LTP). Magnitude and duration of the heterosynaptic LTP were dependent on number, frequency, and intensity of the stimuli. No LTP was produced by a train to the ICN. Heterosynaptic LTP was also observed in the absence of C6. Because of the limited subliminal fringe of the test input under this condition, the LTP was of small magnitude. Heterosynaptic LTP also of the heart rate (HR) response to a test stimulus was observed after a conditioning train. 3. The conditioning train produced a displacement to the right of the dose-response curve for C6. The intravenous dose of C6 required for 50% attenuation of the test CAP increased from 0.84 +/- 0.15 (SE) mg/kg pretrain to 2.56 +/- 0.46 mg/kg posttrain (n = 5, P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

In cats anesthetized with sodium pentobarbital, a supramaximal 40-Hz, 30-s train to the cervical sympathetic trunk, during block of ganglionic cholinergic transmission with hexamethonium and scopolamine, produced a delayed, slow, small amplitude contraction of the nictitating membrane that persisted for several minutes after the end of the stimulus train. The post-stimulus component of the response was due to afterdischarge of the ganglion cells, since section of the post-ganglionic axons at the end of the train resulted in elimination of this component. The amplitude of the slow nictitating membrane response was enhanced in a dose-dependent manner by i.v. injection of naloxone. The enhancement was detectable at a dose as low as 1 μg/kg and was maximal at 10 μg/kg. During continuous preganglionic stimulation at 40 Hz, the amplitude of the slow nictitating membrane response reached a peak in 2–4 min and then faded with time until it became undetectable. Time for 90% decay was 82 ± 5 min (n = 18). The nictitating membrane response to postganglionic nerve stimulation was not modified by prolonged preganglionic stimulation. In three cats, the cervical sympathetic trunk was split into two bundles and one bundle was stimulated continuously at 40 Hz until the slow response disappeared. At this time stimulation of the unconditioned bundle evoked a slow response of normal appearance. This suggests that the process underlying the fade involves only the conditioned axons. Recovery from the fade was slow, the response approaching control by 24 h post-stimulus. In contrast, the nictitating membrane response mediated by ganglionic nicotinic transmission was of amplitude similar to control within the 1st min after the end of a 2-h period of continuous 40 Hz stimulation. The fade and recovery of the slow response may be the consequence of presynaptic events and reflect the exhaustion followed by slow replenishment of the releasable stores of the noncholinergic transmitter. The enhancement of the slow response by naloxone suggests that an inhibitory noncholinergic transmitter, presumably an opioid, is also released by the preganglionic axon terminals.Key words: peptides, opioids, ganglionic transmission, cotransmitters, synapse.

Large dense core vesicles (LDCV), a possible site for neuropeptide storage, were counted within synaptic boutons of the superior cervical ganglion of anaesthetized cats in the unstimulated condition and 2 h, 1 day or 10 days after 40 Hz 2 h stimulation of the cervical sympathetic trunk. The number of LDCV decreased markedly after stimulation. It was lowest at 2 h and then recovered slowly. Ten days post-train the number was close to, but still significantly lower than, control. This time course of recovery is much longer than for agranular vesicles number, which is known to be back to normal within 2 h of the end of a comparable stimulus train. The post-stimulus loss of LDCVs is consistent with the hypothesis that LDCVs are released by the nerve terminal. The slow recovery is consistent with the hypothesis of replenishment by axoplasmic transport.

These experiments investigate the effect of block, by colchicine, of fast axonal transport in the cat's cervical sympathetic trunk (CST) on the superior cervical ganglion's choline acetyltransferase (ChAT) enzyme activity, acetylcholine (ACh) content, and ACh release. Electron microscopy on the segment of the CST exposed to colchicine 1 or 4 days earlier showed disappearance of microtubules and accumulation of vesicles and smooth membrane tubules but no disruption of the axonal cytomatrix. At 4 days following colchicine treatment, the number and size of synaptic boutons per grid square in the ganglion ipsilateral to the colchicine-treated CST were similar to those in the control ganglion. At 2 and 4 days following exposure of the CST to colchicine, ChAT activity in the ipsilateral ganglion was reduced to 78 +/- 8 and 54 +/- 8% of control values, respectively. ACh stores in the ganglia were also reduced (to 81 +/- 6% of control values at 2 days and to 51 +/- 5% of control values at 4 days). Ganglionic transmission and its sensitivity to blockade by hexamethonium during 2-Hz CST stimulation were not impaired at day 4 postcolchicine. ACh release evoked by 2-Hz stimulation of colchicine-treated axons was similar to release from untreated axons, despite the decrease in the ganglionic ACh content. In contrast, ACh release evoked by 20-Hz stimulation was depressed. The amount of ACh released during 5-Hz stimulation in the presence of vesamicol by the terminals of colchicine-treated axons was similar to that released by the terminals of untreated axons. These results suggest the following conclusions: (a) Colchicine-sensitive fast axonal transport contributes significantly to maintaining ChAT stores in preganglionic axon terminals. (b) The half-life of ChAT in sympathetic preganglionic terminals is approximately 4 days. (c) One consequence of colchicine-induced block of axonal transport is a reduced ACh content of preganglionic nerve terminals. (d) This decrease in ACh content appears to be the result of a loss in a reserve transmitter pool, whereas the size of the readily releasable compartment is maintained.

We studied the effect of the adenylate cyclase activator forskolin, of protein kinase C-activating phorbol esters and of prolonged preganglionic input activation on the inhibitory response of the perfused superior cervical ganglion of the cat to exogenous met-enkephalin (Met-ENK). Met-ENK inhibited, in a concentration-dependent manner, the postganglionic compound action potential evoked by cervical sympathetic trunk stimulation. The inhibition was reversible, was blocked by naloxone as well as by pertussis toxin and showed no homologous desensitization in the concentration range 0.01-10 microM. Pretreatment of the ganglion with 4 beta-phorbol 12,13-dibutyrate or 4 beta-phorbol 12,13-diacetate depressed the Met-ENK response for several hours, while pretreatment with forskolin had no effect. This action of phorbol esters was prevented by the protein kinase inhibitor H-7 but not by the calmodulin antagonist W-7 or the protein kinase A inhibitor HA 1004 and was calcium-dependent. Recovery of the response from the depression produced by phorbol esters was not affected by a protein synthesis inhibitor. A 40 Hz 20 min stimulus train to the cervical sympathetic trunk mimicked the effect of phorbol esters, depressing for several hours the inhibition produced by Met-ENK. Stimulus trains of duration shorter than 5 min or frequency lower than 5 Hz were ineffective. This effect of prolonged preganglionic stimulation occurred even when the stimulus train was delivered during complete block of nicotinic and muscarinic ganglionic transmission but was lost when the stimulus train was delivered during perfusion with calcium-free Krebs. The protein kinase inhibitor H-7 prevented the depression of the Met-ENK response by the train, while W-7 and HA 1004 had no effect. These findings suggest that, in the superior cervical ganglion of the cat, a kinase, activated by phorbol esters and inhibited by H-7, exerts a long-term control of the ganglion cell responsiveness to opiate receptor activation. A similar mechanism can be synaptically activated by a non-cholinergic transmitter, released by the preganglionic axons during prolonged, high frequency, activity.

Large dense-cored vesicles are transported centrifugally in the cervical sympathetic trunk and are depleted in a calcium-dependent manner from synaptic boutons of the cat superior cervical ganglion during orthodromic stimulation at 20-40 Hz [P. Weldon et al. (1993) Neuroscience 55, 1045-1054]. In the present study, we tested in awake cats whether the normal tonic firing of the sympathetic preganglionic neuron contributes to the turnover of large dense-cored vesicles in synaptic boutons of the superior cervical ganglion. Tetrodotoxin was applied with a mini-osmotic pump to one cervical sympathetic trunk, while vehicle alone was applied to the contralateral cervical sympathetic trunk, for two, four or seven days. The appearance of Horner syndrome ipsilateral to the tetrodotoxin application demonstrated block of action potential propagation. Both superior cervical ganglia were excised and processed for electron microscopy. The number of large dense-cored vesicles per bouton cross-section was higher in the ganglion with tetrodotoxin-treated input than in the control. The content at four days was higher than at two days; the content at seven days was similar to that at four days. The number of lysosomes per bouton profile also increased in the ganglion with tetrodotoxin-treated input. No changes were observed in size of bouton profiles, number of boutons or of synapses per grid square and length of the presynaptic densities in the ganglion with tetrodotoxin-treated input.(ABSTRACT TRUNCATED AT 250 WORDS)

In anaesthetized cats, a short train to the cervical sympathetic trunk (CST), of frequency between 0.5 and 5.0 Hz, inhibited the postganglionic compound action potential (CAP) evoked by a test shock to the CST. When the CST was split into two bundles, the inhibition was obtained both homosynaptically and heterosynaptically. The inhibition was antagonized by naloxone and enhanced by peptidase inhibitors. Leu-enkephalin (ENK), injected into the arterial supply of the superior cervical ganglion (SCG) depressed the CAP. The ENK-evoked inhibition was antagonized by naloxone, enhanced by peptidase inhibitors and was occluded by the train-evoked inhibition. After prolonged stimulation of the CST (5–20 min at 5 Hz), the inhibition produced by a short train was depressed (faded). The fade was greater the longer the stimulation. After 20 min of stimulation the inhibition was absent. At this time the depressant effect of ENK was unchanged. When only one bundle of the split CST was stimulated at 5 Hz for 20 min, only the inhibition produced by that bundle faded, although both bundles converged onto a common set of ganglion cells. Thus, the fade is not due to receptor desensitization but more likely to failure to release the inhibitory mediator by the overstimulated axons. When, after 20 min of 5 Hz stimulation, no further stimuli were applied, the inhibition recovered in 60 min (fast recovery). However, recovery was slower when stimulation lasted longer than 20 min. Recovery took three days when stimulation lasted 80 min (slow recovery). The inhibition was markedly depressed after application of colchicine to the CST three days earlier. This suggests significant spontaneous release of the endogenous opioid. This treatment also prevented the slow recovery from the fade. The fast recovery from the fade was not prevented by protein synthesis inhibitors nor by a colchicine block of the CST. The data are consistent with the hypothesis that the transmitter mediating the naloxone-sensitive inhibition in the SCG of the cat is present in the preganglionic axon terminals in a small, readily releasable, pool which is exhausted by 20 min stimulation at 5 Hz. A presumably local mechanism, which is exhausted by 80 min of stimulation at 5 Hz, is responsible for the fast recovery. The slow recovery presumably involves protein synthesis and axonal transport.

The morphological and electrophysiological properties of neurons in the intermediolateral nucleus (IML) were studied in the transverse and longitudinal slice of guinea-pig thoracic spinal cord (T2-T3) using intracellular staining and recording techniques. Two morphologically different types of neurons were observed: fusiform cells with craniocaudally oriented dendrites, and multipolar cells with dendrites diffusely extending in the IML. The ratio of fusiform to multipolar cells was 4:1. The fusiform cells were identified as sympathetic preganglionic neurons (SPNs) by their antidromic responses to stimulation of the ventral root exit zone, while the multipolar cells were not antidromically activated by stimulation of this site. Both cell types showed similar resting membrane potential and input resistance. The tonic responses of these neurons to hyperpolarizing current pulses were characteristically different: the SPNs had a marked hyperpolarizing sag at the break of the pulse, caused by an A current, while the unidentified neurons showed no A current. In addition, the SPNs had much longer duration of spike and afterhyperpolarization, as well as lower frequency of spontaneous or current-evoked firing, than the unidentified neurons. These observations suggest that, in the absence of the criterion of antidromic activation by stimulation of the axon, it is still possible to differentiate SPNs from other IML neurons on the basis of morphological and electrophysiological properties of the neuron.

Neostigmine evoked bradycardia in vagotomized, propranolol-treated cats. Heart rate decreased by 50% with 0.4 +/- 0.2 mg/kg (mean +/- S.D.) i.v. of neostigmine. The bradycardia was attenuated after acetylcholine (ACh) depletion in the cardiac parasympathetic pathway suggesting ACh release within this pathway was involved. The bradycardia was unchanged after preganglionic terminal degeneration suggesting ACh release was from cardiac ganglion cells. Edrophonium produced a much weaker bradycardia suggesting the anticholinesterase effect of neostigmine may not produce the bradycardia. The neostigmine-induced bradycardia was blocked by systemic atropine (ED50, 0.005 +/- 0.001 mg/kg), pancuronium bromide (ED50, 0.033 +/- 0.021 mg/kg), pirenzepine (ED50, 74.7 +/- 7.9 micrograms/kg), hexamethonium (ED50, 8.3 +/- 1.6 mg/kg) and d-tubocurarine (ED50, 8.6 +/- 3.0 micrograms/kg). The doses of hexamethonium and d-tubocurarine that blocked the neostigmine-induced bradycardia were significantly higher than required for blocking the bradycardia produced by vagus nerve stimulation. Hexamethonium (60 mg/kg i.v.) had no effect on the bradycardia produced by the muscarinic agonist methacholine (100-300 micrograms/kg/min i.v.). The dose of pirenzepine that blocked the neostigmine-induced bradycardia was lower than required for blocking the bradycardia produced by vagus nerve stimulation. McN-A-343 ([4-hydroxy-2-butynyl]-1-trimethyl ammonium m-chlorocarbanilate chloride) (1 mg/kg i.v.) did not produce bradycardia. These observations suggest neostigmine evokes bradycardia by activation of ACh receptors on cardiac ganglion cells producing ACh release and activation of cardiac M2 receptors. The low sensitivity of the neostigmine-induced bradycardia to pirenzepine, and the failure of McN-A-343 to evoke bradycardia, suggest the receptor on cardiac ganglion cells is not an M1-type.(ABSTRACT TRUNCATED AT 250 WORDS)

The relation between arterial O2 tension (PaO2) and the firing rate of sympathetic preganglionic neurons (SPN) was studied in 16 strands of the cervical sympathetic trunk (CST) during graded isocapnic hypoxia in 11 sinoaortic-denervated central nervous system (CNS)-intact, anesthetized cats. SPN firing rate was independent of PaO2 from normoxia down to a PaO2 of 40 Torr. Below this PaO2 level three response patterns were observed, i.e., an excitatory response (n = 8), a depressant response (n = 3), and a mixed response consisting of a depression of firing at less severe hypoxic levels and an increase in firing rate at more extreme hypoxic levels (n = 5). Similar response patterns were also observed in four strands of the CST in three unanesthetized, sinoaortic-denervated, midcollicular decerebrate preparations. Systemic arterial pressure decreased in all cats as PaO2 decreased. Phrenic nerve activity also decreased in all cats with a course resembling that of the depression of sympathetic firing and disappeared at a PaO2 of 20 Torr. The data suggest that systemic hypoxia in the sinoaortic-denervated, CNS-intact or decerebrate animal activates both excitatory and depressant mechanisms acting on SPNs.

The electrical activity of phrenic and recurrent laryngeal motoneurons was recorded during activation of urinary bladder afferents in pentobarbital-anesthetized, paralyzed, artificially ventilated cats. Distension or spontaneous neurogenic contraction of the bladder decreased the amplitude and frequency (bursts/min) of the burst activity in these two nerves. The decrease in frequency was due mainly to a prolongation of the interburst interval (duration of expiration). Similar effects were observed during low-frequency electrical stimulation of pelvic nerve afferents with conduction velocities in the A gamma-A delta range. High-frequency stimulation of pelvic nerve afferents increased the amplitude of the burst activity in these two nerves. These results suggest an action of bladder afferents on the brain stem oscillator controlling respiratory frequency and drive. Some of the excitatory and inhibitory effects of bladder afferents on phrenic motoneurons may be mediated through spinal circuits.

The relation between arterial O2 tension (PaO2) and the firing rate of sympathetic preganglionic neurons (SPN) of the cervical sympathetic trunk was studied during graded isocapnic hypoxia and hyperoxia in unanesthetized acute C-1 spinal cats. In the PaO2 range between 40 and 400 Torr there was no relation between the two variables. Below 40 Torr firing rate increased as PaO2 decreased, reaching an average peak value of ten times control at a PaO2 of 20 Torr. Mean arterial blood pressure (MABP) was also independent of PaO2 between 40 and 400 Torr and increased by an average of 25% at PaO2 values below 40 Torr. Intravenous administration of hexamethonium or phentolamine abolished the MABP response but not the SPN response to hypoxia. Pentobarbital (5-60 mg/kg iv) did not modify the SPN response to hypoxia, although normoxic SPN background firing was considerably depressed. The excitatory effect of hypoxia seems independent of excitatory afferent input and appears to be a general property of SPN.

In the in vitro slice preparation of upper thoracic cord of the cat, noradrenaline, at concentrations of 10-50 microM, induced rhythmic bursting in 30% of sympathetic preganglionic neurons of the intermediolateral nucleus. The bursting frequencies were between 0.2 and 1.0 Hz at membrane potentials between -45 and -65 mV. The bursting rhythm could be reset by short intracellular current pulses. In the presence of tetrodotoxin noradrenaline produced a rhythmic oscillation in membrane potential in the same frequency range as the bursting. The frequency of oscillation was voltage dependent. Neuronal input resistance decreased at the oscillation peak and the oscillation was abolished by cobalt or low calcium.

The effects on the pressor response to cerebral ischemia (CIR) of superfusion of the ventral medullary surface with artificial cerebrospinal fluid (CSF) containing local anesthetic was investigated in 10 sinoaortic-denervated, anesthetized cats. Prior to application of the local anesthetic, occlusion of the common carotid and vertebral arteries caused an increase in mean systemic arterial pressure (SAP) of 58 +/- 7 mmHg (+/- SE) from an initial level of 98 +/- 6 mmHg. Following 108 +/- 15 s of superfusion with artificial CSF containing 2% procaine, the CIR decreased to 19 +/- 3 mmHg. At this time phrenic nerve activity had been eliminated but basal SAP had only decreased by 14 +/- 2 mmHg, and significant neurogenic vasomotor tone remained. The residual CIR can be accounted for by the passive increase in systemic resistance due to occlusion of the cerebral vascular bed. The effects of procaine were reversible. On attenuation of the CIR, electrical stimulation of pressor points 2–4 mm from the ventral medullary surface was still effective. Autoradiographic analysis following application of 14C-labeled lidocaine showed that attenuation of the CIR occurred when estimated concentrations of the anesthetic sufficient to block nerve conduction extended 85 micron from the ventral medullary surface. These results indicate that the CIR is mediated by superficial structures in the ventral medulla that are not involved in the generation of a major fraction of basal vasomotor tone.

Focal electrical stimulation of the slice of the cat thoracic cord evoked in sympathetic preganglionic neurons a slow inhibitory postsynaptic potential (IPSP) associated with decreased neuronal input resistance. The slow IPSP decreased in amplitude with membrane hyperpolarization and reversed at about -90 mV. It increased in amplitude in low potassium and decreased in high potassium. Noradrenaline (NA) at doses of 10-50 microM caused in some of these cells a hyperpolarization with properties similar to those of the slow IPSP. Both the slow IPSP and the NA-evoked hyperpolarization were abolished by yohimbine, but not by prazosin or propranolol. These data suggest that both responses are due to an increase in potassium conductance and that NA may be the mediator of the slow IPSP evoked by focal stimulation.

The properties of sympathetic preganglionic neurone activity during expiration were studied in pentobarbitone-anaesthetized (n = 26) and in non-anaesthetized, mid-collicular decerebrate (n = 5), paralysed, artificially ventilated cats in which the electrical activity of the phrenic nerve and of the cervical sympathetic trunk was recorded. In control conditions (end-tidal PCO2 between 35 and 40 mmHg, zero end-expiratory pressure) sympathetic activity during expiration was either steady at a low level (n = 11) or showed a modest progressive increase from a low level in early expiration (n = 17). Very infrequently (n = 3), it showed a transient increase during the second half of expiration. Artificial ventilation with positive end-expiratory pressures in the range from 2.1 +/- 0.4 (mean +/- S.D.) to 6.7 +/- 0.6 cmH2O caused, in cats with intact vagus nerves, an increase in sympathetic neurone activity during the second half of expiration. Within this range of pressures, the magnitude of the increase was related to the magnitude of the positive end-expiratory pressure. This effect reversed at higher positive end-expiratory pressures. Pressures in excess of 10.2 +/- 1.8 cmH2O caused inhibition of sympathetic activity. The sympatho-excitatory effect of positive end-expiratory pressure disappeared after bilateral cervical vagotomy. With intact vagus nerves, it also disappeared at levels of systemic hypocapnia (end-tidal PCO2 less than or equal to 15 mmHg) which abolished phrenic nerve activity. In hypocapnia, artificial ventilation with peak tracheal pressures greater than 7.2 +/- 1.1 cmH2O caused inhibition of sympathetic activity, while ventilation with lower end-expiratory pressures had no effect on sympathetic activity. It may be concluded that the sympatho-excitatory effect of positive end-expiratory pressure is mediated by vagal afferents and requires a certain level of brain-stem respiratory neurone activity. Sympatho-excitation during expiration was also observed, in normocapnic conditions, during short-duration static lung inflation with tracheal pressures in the range from 2.5 +/- 0.3 to 7.0 +/- 0.8 cmH2O as well as during artificial ventilation with zero end-expiratory pressure when lung inflation occurred in expiration. These responses were abolished by bilateral cervical vagotomy and during systemic hypocapnia. Sympatho-excitation during expiration was also observed when systemic hypercapnia was produced in vagotomized cats by artificial ventilation with gas mixtures containing 5 or 10% CO2.(ABSTRACT TRUNCATED AT 400 WORDS)

Cardiac output and blood flow to the kidney, hind limb, and splanchnic area were studied in anesthetized dogs during the arterial pressure rise due to carotid occlusion. All the vascular beds studied exhibited an increased resistance to flow. Cardiac output was maintained constant through an increased cardiac work in spite of the increased arterial pressure. This increased cardiac work was reflexly produced as evidenced by the action of the adrenergic blocking agent dichloroisoproterenol (DCI). This drug allowed the reflex vasoconstriction and the consequent pressure rise, but preventing the reflex chronotropic and inotropic sympathetic effects on the heart resulted in a fall of cardiac output during carotid occlusion. It is concluded a) that the reflex increase in vasomotor tone is the main factor in the pressor response to carotid occlusion and b) that the reflex effect on the myocardium is an important factor in regulating cardiac output in this situation.

Publisher Summary This chapter summarizes the results obtained in an analysis of catecholamines (CA) actions on sympathetic preganglionic neurons (SPNs) carried out by intracellular recording in the slice of the upper thoracic spinal cord of the cat. These data describe the actual action of CA on the SPN, and therefore permit the prediction of the effects that CA are likely to mediate in the intact animal. Several lines of evidence suggest that CA have a role as mediators, or modifiers, of synaptic actions on the SPN. The intermediolateral (IML) nucleus of the gray matter of the thoracic spinal cord is rich in CA-containing axons. Several CA-synthesizing neuron sets of brainstem and hypothalamus, some of which are located in the ventrolateral medulla, are considered as the source of this dense CA innervation of the IML column. CA-containing axons form typical synapses with identified SPNs. Binding studies show that adrenoceptors are present in the IML nucleus. The iontophoresis of CA on SPNs produces depressant effects mediated by a α 2 -adrenoreceptor.

In cats anesthetized with chloralose-urethan, vagotomized, paralyzed, and artifically ventilated, superficial radial (cutaneous) and hamstring (muscle) nerve afferents were stimulated while phrenic nerve electrical activity was recorded. The results obtained with both types of nerves were similar. Stimulation in mid and late expiration advanced the onset of the next inspiration, shortening its duration. Stimulation in early inspiration advanced, while that in late inspiration delayed, the onset of the next expiration. These effects were often accompanied by changes in phrenic motoneuron firing patterns (earlier recruitment, increased discharge frequency, increased slope of integrated phrenic neurogram). Repetitive somatic afferent stimulation produced sustained increases in respiratory frequency in all cats and in half of them entrainment of respiratory frequency to the frequency of stimulation occurred at ratios such as 4:3, 4:5, 1:2, 1:3, 1:4, and 1:7. The lowest stimulus intensity required for evoking these phase shifts was between 5 and 10T (threshold of most excitable fibers) for muscle afferents and between 1 and 2T for cutaneous afferents. These results demonstrate the existence of a reflex mechanism capable of locking respiratory frequency to that of a periodic somatic afferent input. They also provide an experimental basis for the hypothesis that reflexes are resposible for the observed locking between step or pedal frequency and respiratory rate during exercise in man.

Intracellular recordings were made from sympathetic preganglionic neurons of the lateral horn in slices of cat thoracic cord maintained in vitro. Focal electrical stimulation of the slice evoked, in addition to the already described fast EPSPs, EPSPs of several seconds duration. The slow EPSPs, like the fast EPSPs, were graded with stimulus intensity and were abolished by TTX or low Ca and high Mg superfusion. The slow EPSP decreased in amplitude with membrane hyperpolarization and was nullified at -90 mV but did not reverse with further hyperpolarization. The slow EPSP was abolished by phentolamine or prazosin but not by yohimbine. Noradrenaline NA, 10-50 microM) caused in 30% of neurons a TTX-resistant depolarization. The NA-evoked depolarization had the same characteristics as the slow EPSP with respect to sensitivity to membrane potential and to adrenergic blockers. These results suggest that NA, acting on an alpha 1-receptor, may be the mediator of the slow EPSP evoked in this neuron by focal stimulation.

The aim of the present study was to evaluate the organization of neural circuitry responsible for the intersegmental transmission of input from urinary bladder afferents to sympathetic preganglionic neurons (SPNs). The electrical activity of SPNs was recorded from axons of the cervical sympathetic trunk in anesthetized central nervous system (CNS)-intact and in unanesthetized midcollicular-decerebrate or acute C1 spinal cats. In all three preparations, tonically active SPNs were excited or inhibited by 1) electrical stimulation of myelinated afferents of the pelvic or hypogastric nerve, both of which contain bladder afferents, and 2) spontaneous contraction or distension of the urinary bladder. The SPN responses to bladder distension were abolished by pelvic nerve section. A comparison of responses of SPNs in CNS-intact and acute spinal animals to electrical stimulation of pelvic nerve afferents suggests that both propriospinal and supraspinal circuits are involved in the intersegmental transmission of input from bladder afferents to SPNs.

Intracellular recordings were made from sympathetic preganglionic neurons of the intermediolateral nucleus, in the slice of the T3 segment of the cat spinal cord. Depolarization to -60 mV from membrane potentials more negative than -75 to -80 mV caused a transient outward rectification which decayed in 0.5-2.0s. The rectification was sensitive to changes in K concentration, was abolished by 4AP (2 mM), and was unaffected by low Ca (0.25 mM), Co (2 mM), Ba (1 mM), TEA (20 mM) or intracellular Cs. These properties suggest that the rectification is due to a transient K-current similar to the A-current described in other neurons.

The effect on heart rate of close i.a. injection of neurotensin (NT), substance P (SP), and vasoactive intestinal peptide (VIP) into the decentralized right stellate ganglion was tested in anaesthetized spinal cats. These peptides are present in the stellate ganglion and may mediate the stellate ganglion cell excitation underlying a previously described slow cardioacceleration evoked by preganglionic stimulation during block of cholinergic transmission. NT (Tyr11-NT) at doses of 25–200 μg produced increases in heart rate of 10–125 beats/min (bpm) and of slow time course. At the dose of 100 μg, NT produced a cardioacceleration of 56 ± 8.4 bpm(mean ± SEM, n = 13) with an onset latency of 23 ± 4 s, a slow rise to peak (rise time 62 ± 4.5 s), and a half decay of 167 ± 14 s. A cardioacceleration of comparable magnitude (78 ± 3.8 bpm) caused by close i.a. administration of acetylcholine (100 μg, n = 13) had an onset latency of 2 ± 1 s, a fast rise to a sharp peak (rise time 3 ± 1 s), and a half decay of 23 ± 4 s. The analogues, Phe11-NT and Trp11-NT, as well as the stereoisomer, D-Tyr11-NT, had no effect on heart rate when injected at doses up to 400 μg. The NT-evoked cardioacceleration was blocked by propranolol or by section of the inferior cardiac nerve and may therefore be attributed to prolonged excitation of stellate ganglion cells. Administration of hexamethonium and atropine was without effect on the NT response. Intravenous injection of NT at the same doses used by intra-arterial injection (25–200 μg) was without appreciable effect on heart rate. At doses of up to 400 μg by close i.a. injection, VIP and SP had no effect on heart rate. Two observations suggest that NT is a likely candidate as mediator of noncholinergic excitation of stellate ganglion cells. One is the finding, reported here, of a similarly slow time course for the NT-evoked cardioacceleration and for the slow cardioacceleration evoked by preganglionic stimulation during block of cholinergic transmission. The other is the previously reported finding that NT-like immunoreactivity in the stellate ganglion is depleted by prolonged stimulation causing loss of the neurally evoked slow cardioacceleration.

The background discharge of sympathetic preganglionic neurones shows a marked inspiration-synchronous component which is known to originate from within the central nervous system. The contribution of this component to total neurogenic vasoconstrictor tone is unknown. In order to estimate its extent we have exploited the inspiration-suppressing effect of a group of low threshold afferent fibres in the superior laryngeal nerve. The electrical activities of the cervical sympathetic trunk and of the phrenic nerve were recorded in pentobarbitone-anaesthetized, paralysed, artificially ventilated, sino-aortic denervated and vagotomized cats, together with the perfusion pressure of an innervated hind limb perfused at a constant flow rate. Repetitive stimulation of the superior laryngeal nerve at an intensity just sufficient to suppress phrenic nerve activity inhibited the inspiration-synchronous sympathetic discharge and caused hind limb vasodilatation. This vasodilatation was abolished by hexamethonium or phentolamine, but was not affected by atropine or propranolol. Following the elimination of phrenic nerve activity and inspiration-synchronous sympathetic discharge by systemic hypocapnia, repetitive stimulation of the superior laryngeal nerve either failed to affect the residual sympathetic activity and hind limb perfusion pressure, or caused an increase of both. Stimulation of the superior laryngeal nerve with short (0.2 s) trains of stimuli, delivered at selected times of the respiratory cycle for several consecutive cycles, had similar effects on phrenic nerve bursts, inspiration-synchronous sympathetic discharge and hind limb perfusion pressure. Stimulation at progressively earlier times during inspiration produced a graded reduction in all three variables, while stimulation during late inspiration or early expiration had no effect on any of them. The results suggest that the vasodilator reflex, elicited by inspiration-suppressing afferents in the superior laryngeal nerve, results from selective abolition of the excitatory input which causes the inspiration-synchronous discharge of sympathetic neurones. The magnitude of the hind limb vasodilatation can therefore be taken as an indication of the extent of control of hind limb vasoconstrictor tone exerted by this particular input. By comparing the magnitude of the reflexly evoked vasodilatation with that of the vasodilatation resulting from ganglionic blockade, it was estimated that 24.2% of the neurogenic vasoconstrictor tone of the hind limb was attributable to the inspiration-synchronous component of sympathetic discharge.

Sympathetic preganglionic neuron (SPN) activity (mass or single unit discharge) was recorded extracellularly from axons of the cervical sympathetic trunk (CST) in unanaesthetized, midcollicular decerebrate or C1 spinal anemically decerebrate cats which were paralyzed and artificially ventilated. Phrenic motoneuron activity was also recorded in the midcollicular preparation. The drug 4-aminopyridine (4-AP) was administered intravenously. Doses of 0.5 to 1.0 mg/kg increased the background firing rate of SPNs in a dose-dependent manner. These doses increased the repetition rate of phrenic nerve bursts and, in a parallel fashion, that of the inspiration-synchronous activity of the CST. In contrast, the action of 4-AP on activity evoked by stimulation of high threshold myelinated afferents in the radial, femoral or pelvic nerve, or in the L7 dorsal root, was very variable. The evoked responses could be increased, decreased or not affected by the drug. Thus, treatment with 4-AP produces a graded increase in background firing of SPNs, but the drug is ineffective at reliably enhancing somato- or viscero-sympathetic reflexes.