The location and properties of preganglionic vagal cardiomotor neurones in the rabbit

The origins of preganglionic vagal neurones which slow the heart in the rabbit have been examined with standard neurophysiological stimulation and recording techniques. The activity of 216 neurones projecting to the right cervical vagus nerve have been recorded in localized areas of the brain stem. Thirty-six of these neurones were classified as cardiomotor neurones since they had properties similar to those described for such neurones in the cat. All had efferent axons in the range of B fibers. They could be synaptically activated by electrical stimulation of the ipsilateral aortic nerve which in the rabbit is solely barosensory. The majority of these neurones (70%) were spontaneously active and those which were normally silent could be made to fire by iontophoretic application ofdl-homocysteic acid (an excitant amino acid). This spontaneous, or evoked, activity showed evidence of a pulse rhythm (of baroreceptor origin) and respiratory modulation (firing predominantly during expiration). In response to application ofdl-homocysteic acid, the neuronal excitation was usually accompanied by a small but significant bradycardia. Histological examination showed that these neurones were located in both the dorsal vagal nucleus and the nucleus ambiguus.     

Age-related loss of cardiac vagal preganglionic neurones in spontaneously hypertensive rats

Despite the findings that impaired vagal control of the heart rate occurs in human hypertension, leading to greater cardiovascular risk, the mechanism of this impairment is as yet unknown. Observations in humans and experiments in the spontaneously hypertensive rat (SHR) suggested that such impairment may be related to an anomaly in central vagal neurones. We therefore set out to determine whether the numbers and distribution of cardiac-projecting vagal preganglionic neurones in the medulla of adult (12 week) hypertensive SHR are different from those in young (4 week) prehypertensive SHR and in age-matched Wistar-Kyoto (WKY) rats of two age groups. The number of vagal neurones, identified by labelling with the fluorescent tracer DiI applied to the heart, was essentially similar in the three areas of the medulla analysed (dorsal vagal nucleus, nucleus ambiguus and intermediate reticular zone) in young SHR and young or adult WKY rats. In contrast, fewer vagal neurones were labelled in adult SHR compared with young SHR or WKY rats. This difference was due to highly significant reductions in vagal neurones in the dorsal vagal nucleus and nucleus ambiguus on the right side of the medulla. These observations suggest that a loss of parasympathetic preganglionic neurones supplying the heart with axons in the right vagus nerve, or a remodelling of their cardiac projections, may explain the known impairment of the baroreceptor reflex gain controlling heart rate in hypertension.     

Discharge patterns of preganglionic neurones with axons in a cardiac vagal branch in the rat

The fibre types that run in a vagal branch projecting to the rat heart are described in this study. In order to obtain spontaneous discharge in this vagal branch and optimal recording conditions, we compared the decerebrate state to urethane, urethane-chloralose and pentobarbital-chloralose anaesthesia with regard to level of chronotropic cardiac vagal tone. Administration of atropine (2 mg kg(-1), I.V.) significantly decreased baseline cardiac interval only in the decerebrate and urethane-anaesthetised rat (by 0.018 +/- 0.001 s and 0.019 +/- 0.002 s, respectively). As a result of these experiments, urethane was chosen as the anaesthetic for all subsequent studies. Using a heart rate signal-averaging method we demonstrated that rat cardiac vagal preganglionic neurones innervating the sinoatrial node should have an expiratory discharge pattern, as reported in other species. However, only 5 % of chronotropic vagal tone was found to be subject to respiratory sinus arrhythmia. A suction microelectrode method, combined with spike-triggered averaging, was employed to record activity from a total of 58 vagal afferents that had axons in this branch. Approximately 75 % of these latter sensory fibres displayed cardiac rhythm. In a separate study we also recorded 318 preganglionic neurones with axons in the right cardiac vagal branch of the rat. Respiratory-modulated preganglionic units were statistically less common than tonically firing units. Six preganglionic subtypes were categorised according to conduction velocity and respiratory discharge pattern. Myelinated B-fibre and unmyelinated C-fibre types were found to be equally prevalent and equally likely to be reflexly excited during the pulmonary chemoreflex and the peripheral arterial chemoreflexes. The electrophysiological analysis has shown how diverse the discharge patterns of the preganglionic neurones or interneurones are whose axons course in the right cardiac vagal branch of the rat. The results of these experiments demonstrate the usefulness of combining spike discrimination with multiple spike-triggered averaging to simultaneously record B and C centrifugal vagal efferents.     

Fibroblast growth factor-1 improves the survival and regeneration of rat vagal preganglionic neurones following axon injury

Self-initiated leg movement in standing humans is preceded by a medio-lateral preparatory balance adjustment (PBA); however, such preparatory balance control is often absent in reflex-like stepping responses evoked by whole-body instability. The presence or absence of the PBA may reflect a task-dependent modulation of the response serving to preserve lateral stability (PBA present) or avoid delay in the lifting of the foot (PBA absent). To examine whether such task-dependent modulation can occur during more stereotypical limb movements, we examined spinally-mediated withdrawal responses evoked by noxious stimulation of the foot. Results showed that rapid limb withdrawal was preceded by a large PBA when subjects were standing but not when they were supine. The PBA caused limb withdrawal to the noxious stimulation to be delayed. However, the onset of the PBA in the standing trials was equivalent in timing to the onset latency of the classic withdrawal responses recorded during the supine trials. Evidence of a preparatory balance adjustment evoked, in advance of a delayed withdrawal response, at very rapid latencies (underlying muscle activation at 70-120 ms) may raise new questions about the neural mechanisms underlying the co-ordination of balance and movement.     

Swallow-evoked action potentials in vagal preganglionic efferents

Swallow-evoked potentials in the preganglionic vagal fibers were studied using the single-fiber recording technique in anesthetized opossums. Swallows were evoked by tactile pharyngeal stimulation or electrical stimulation of the cut central end of the superior laryngeal nerve (SLN). Swallowing activity was recorded by the mylohyoid electromyogram and esophageal motility. Sixty-six fibers were studied in which swallowing evoked action potentials. The latencies (from the onset of mylohyoid activity) of evoked responses in different fibers varied from 100 ms to 5 s. The discharge rate of the evoked response was 3-8 action potentials per burst. Each burst lasted 1.1 +/- 0.02 (SE)s. The latencies of evoked spike bursts showed a bimodal distribution. In 34 fibers the latencies were less than 1 s, and in 32 fibers the latencies ranged between 1 and 5 s; these are the short- and long-latency fibers, respectively. Short-latency fibers could easily be distinguished from long-latency fibers based on the influence of SLN-stimulus frequency. Short-latency discharges had low thresholds of activation and were sensitive to changes in the frequency of SLN stimulation, since their latencies decreased and their discharge rate increased with increasing SLN-stimulus frequency. On the other hand, the latencies and discharge rates of long-latency discharges were not modified with changing SLN stimulus frequencies. The conduction velocities of 6 short- and 9 long-latency fibers were 5.64 +/- 0.12 and 5.78 +/- 0.12 (SE) m/s, respectively (P greater than 0.05). The relationship between the latencies of swallow-evoked discharges in the short- and long-latency fibers and the esophageal smooth muscle responses suggested that the short-latency discharges may correlate with the latency of initial inhibition, and the long-latency fibers may correlate with latencies of peristaltic contractions. Based on these temporal relationships, we speculate that vagal efferent fibers showing swallow-evoked, short-latency discharges make contact with intramural inhibitory neurons. They may mediate deglutitive inhibition in the body of the esophagus, relaxation of the lower esophageal sphincter, and receptive relaxation of the fundus of the stomach. The fibers showing late discharges make contact with intramural excitatory neurons and participate in their sequential activation. This dual pathway of activation may be responsible for physiological esophageal peristalsis.     

The activity characteristics of the preganglionic pupilloconstrictor neurones

1. Recordings have been made of pupilloconstrictor unit activity within the small-celled component of the oculomotor nucleus of the cat. The identity of these neurones was determined by testing unit responses to a sequence of stimuli which produced dilation and constriction of the sympathectomized pupil.2. The frequency range of unit activity was from 0 to 28/sec. A maximal constriction of the pupil was maintained by unit activity in the region of 8/sec. The resting level of unit activity under chloralose anaesthesia was typically in the range 6-10/sec.3. Stimulation in the defence area of the hypothalamus produced a maximal pupillodilation associated with a complete inhibition of unit activity and a block of the light reflex response.4. Onset of retinal illumination produced a burst discharge followed by a transient silent period and then an over-all increase in the rate of activity.5. Cessation of retinal illumination produced a post-excitatory depression of unit activity lasting from 190 to 700 msec. It is suggested that this effect is produced by retinal afferents activated by light ;off' that exert an inhibitory influence on the pupilloconstrictor neurones.     

Effects of afferent volleys from the limbs on the discharge patterns of interpositus neurones in cats anaesthetized with alpha-chloralose.

1. In cats anaesthetized with alpha-chloralose, micro-electrodes have been used to record the discharge patterns of single neurones in the region of the nucleus interpositus. 2. Almost all cells tested could be antidromically invaded following electrical stimulation of the contralateral red nucleus, showing that they were cerebellar efferent neurones. 3. A little over half of the interpositus neurones were spontaneously active, usually at rates of less than 20 impulses/sec. 4. About 40% of the cells had no spontaneous activity, although they gave brisk responses to electrical stimulation of cutaneous nerves. Such silent units were encountered most frequently in the earlier stages of an experiment, but a number were found more than 15 hr after the beginning of an experiment. 5. Stimulation of cutaneous and mixed nerves of the fore and hind limbs provoked impulse discharges of the cells and also produced phases of deceleration of the resting discharge of spontaneously firing cells. 6. The typical response of an interpositus neurone consisted of a short latency (6-35 msec) discharge, usually separated from a long latency (50-500 msec) discharge by a period of inhibition or return to the resting discharge rate. The two phases of excitation appeared to be independently generated, since in a number of cells one phase appeared without the other. In addition, the later phase of excitation was abolished in all cells tested by a small dose of pentobarbitone which produced very little effect on the earlier phase. The long latency response was quantitatively much greater, sometimes consisting of 50 or more impulses in a response which lasted several hundred msec, but was very variable from one trial to another. 7. The long latency discharge and sometimes the preceding inhibition could readily be mimicked by single shock stimulation of the region of the contralateral inferior olive. Short latency discharges were, however, rarely evoked by olivary stimulation. 8. It is suggested that the short latency responses of the interpositus neurones were a result of synaptic excitation via cerebellar afferents, while the ensuing inhibition was a result of post-synaptic inhibition resulting from the Purkinje cell excitation due to the afferent volleys. It is suggested that the long latency excitation is due to the afferent volleys. It is suggested that the long latency excitation is due at least in part to disinhibition resulting from long pauses in Purkinje cell firing following their activation by climbing fibre afferents. 9. The possibility that these long latency responses have a physiological significance in relation to locomotion is discussed.     

Effects of substance P on sympathetic preganglionic neurones

Antidromically identified sympathetic preganglionic neurones, located in the second thoracic segment of the rat spinal cord, were tested for their response to iontophoretically applied substance P. Substance P increased the firing rate of both 'spontaneously' active and glutamate-activated neurones. As substance P-like immunoreactivity is present in the intermediolateral cell column of the thoracic spinal cord, it is concluded that substance P may be an excitatory transmitter or modulator involved in mediating excitatory drive to sympathetic preganglionic neurones.     

Gastric blood flow in anaesthetized cats

1. The gastric mucosal blood flow has been measured by the amidopyrine clearance technique in anaesthetized cats. The total gastric blood flow has been (a) measured directly and (b) calculated by the Fick principle from the amidopyrine concentrations in gastric arterial and venous blood and the gastric output of amidopyrine.2. Observations on the recovery of added amidopyrine from arterial and venous blood and plasma, and on the rate of transfer of amidopyrine from corpuscles to plasma, support the underlying assumptions of the amidopyrine method of measuring mucosal blood flow.3. If acid solutions are instilled into the stomach the mucosal blood flow of the non-secreting stomach may be measured by the amidopyrine technique.4. Total gastric and mucosal blood flow increased linearly with increase in H(+) secretion, stimulated by histamine or gastrin. The increase in total flow was entirely due to the increase in mucosal flow.5. The relationship between mucosal blood flow and H(+) secretion was the same for histamine and gastrin responses, and was unaffected by maintaining the total flow at a constant level, or by reduction of the circulating blood volume. Increase in blood volume altered the relationship so that there was a greater increment in mucosal blood flow for any increase in H(+) secretion.6. It is concluded that valid observations may be made on secretion and blood flow relationships in acute anaesthetized preparations.     

The identification and characteristics of sacral parasympathetic preganglionic neurones

1. Sacral parasympathetic preganglionic neurones were identified by intracellular and extracellular micro-electrode recording of antidromic potentials in response to stimulation of the pelvic nerve or the second or third sacral ventral roots.

2. The segmental distribution of autonomic neurones varied in different cats. In some cats they were mainly in S2 segment, in others in S3 and in the remainder, in both S2 and S3.

3. The antidromic potentials showed initial segment-somadendritic (IS-SD) inflexions and delayed depolarizations and were slightly less prolonged than those of sympathetic neurones but more prolonged than those of spinal motoneurones. After-hyperpolarization was observed after the antidromic spike potential.

4. The conduction velocities for sacral parasympathetic preganglionic fibres were less than 12·5 m/sec and thus were similar to those of sympathetic preganglionic fibres.

5. Parasympathetic neurones were not excited by micro-electro-phoretically applied 5-hydroxytryptamine, noradrenaline or acetylcholine.

     

The location of cardiac vagal preganglionic motoneurones in the medulla of the cat.

1. Electrophysiological techniques have been used to locate the origin of preganglionic vagal motoneurones supplying the heart of the cat. 2. The right cardiac vagal branches were identified anatomically and their ability to slow the heart was assessed by electrical stimulation. Control experiments revealed that contamination of cardiac branches by bronchomotor and oesophageal efferent fibres was likely to be small. 3. Fifty-seven neurones in the medulla were activated antidromically on stimulating the cardiac branches at up to 5 times the threshold for cardiac slowing. They had axons with conduction velocities between 3 and 15 m/sec, corresponding to B fibres. 4. None of these were located in the region of the dorsal motor nucleus of the vagus, in spite of repeated sampling there, but all were located in the region of the nucleus ambigus. Histological examination of marked neurones (forty-six of the fifty-seven neurones) revealed that they were associated with its principal column, rostral to the obex. 5. Sampling motoneurones of the dorsal motor nucleus revealed that most sent axons down the thoracic vagus below the cardiac branches. Only three of thirty-three could be activated antidromically by high intensity stimulation of the cardiac branches, but on the basis of their thresholds and conduction velocities, it is argued that they were unlikely to be cardio-inhibitory neurones. 6. It is concluded that preganglionic cardio-inhibitory neurones arise not in the dorsal motor nucleus, but in the principal column of the nucleus ambiguus.     

Plateau potentials and membrane oscillations in parasympathetic preganglionic neurones and intermediolateral neurones in the rat lumbosacral spinal cord

Whole-cell patch recordings were made from parasympathetic preganglionic neurones (P-PGNs) and unidentified intermediolateral (IML) neurones in thick slices of the lower lumbar and sacral spinal cord of 14- to 21-day-old rats. The P-PGNs and IML neurones examined were similar in terms of soma sizes, input resistance and capacitance, and displayed a sag conductance as well as rebound firing. In the absence of drugs, the neurones responded with either tonic or adapting firing to depolarizing current steps. However, in the presence of the group I metabotropic glutamate receptor agonist ( RS )-3,5-dihydroxyphenylglycine (DHPG), almost half of the neurones displayed accelerating firing rates during the constant current injection, followed by a sustained after-discharge. In the presence of TTX, plateau potentials were observed. The firing changes and plateaux were blocked by nifedipine, an L-type Ca²⁺ channel blocker, and ( S )-(−)-Bay K8644 was able to produce these firing changes and plateaux in the absence of DHPG, demonstrating the involvement of an L-type Ca²⁺ conductance. Ca²⁺-activated nonspecific cationic conductances also appear to contribute to the firing changes. A few neurones displayed membrane oscillations and burst firing in the presence of DHPG. The results suggest that the firing characteristics of both P-PGNs and other neurones likely to be involved in caudal spinal reflex control are not static but, rather, quite dynamic and under metabotropic glutamate receptor modulatory control. Such changes in firing patterns may be involved in normal pelvic parasympathetic reflex function during micturition, defaecation and sexual reflexes, and may contribute to the abnormal output patterns seen with loss of descending brainstem input and visceral or perineal sensory disturbances.     

Population spatiotemporal dynamics of spinal intermediate zone interneurons during air-stepping in adult spinal cats

The lumbar spinal cord circuitry can autonomously generate locomotion, but it remains to be determined which types of neurons constitute the locomotor generator and how their population activity is organized spatially in the mammalian spinal cord. In this study, we investigated the spatiotemporal dynamics of the spinal interneuronal population activity in the intermediate zone of the adult mammalian cord. Segmental interneuronal population activity was examined via multiunit activity (MUA) during air-stepping initiated by perineal stimulation in subchronic spinal cats. In contrast to single-unit activity, MUA provides a continuous measure of neuronal activity within a ～100-μm volume around the recording electrode. MUA was recorded during air-stepping, along with hindlimb muscle activity, from segments L3 to L7 with two multichannel electrode arrays placed into the left and right hemicord intermediate zones (lamina V-VII). The phasic modulation and spatial organization of MUA dynamics were examined in relation to the locomotor cycle. Our results show that segmental population activity is modulated with respect to the ipsilateral step cycle during air-stepping, with maximal activity occurring near the ipsilateral swing to stance transition period. The phase difference between the population activity within the left and right hemicords was also found to correlate to the left-right alternation of the step cycle. Furthermore, examination of MUA throughout the rostrocaudal extent showed no differences in population dynamics between segmental levels, suggesting that the spinal interneurons targeted in this study may operate as part of a distributed "clock" mechanism rather than a rostrocaudal oscillation as seen with motoneuronal activity.     

Phenotypic traits of the hypothalamic PVN cells innervating airway-related vagal preganglionic neurons

The paraventricular nucleus of the hypothalamus (PVN) integrates multiple inputs via projections from arginine vasopressin (AVP)- and oxytocin (OXT)-containing neurons to the brain stem and spinal cord as well as regulates respiratory and cardiovascular stress-related responses, which also affect airway function. In the present study, we used immunocytochemistry and the retrograde transneuronal tracer, Bartha strain of pseudorabies virus expressing green fluorescent protein (PRV-GFP), to localize AVP- and OXT-producing neurons that project to airway-related vagal preganglionic neurons (AVPNs) innervating intrapulmonary airways. PRV-GFP was microinjected into the upper right lung lobe, and after 4 days survival, hypothalamic tissue sections were processed for co-expression of PRV-GFP and AVP or PRV-GFP and OXT. In addition, in a separate group of five rats, Phaseolus vulgaris leucoagglutinin (PHAL), an anterograde tracer, was injected unilaterally into the PVN and cholera toxin beta subunit was microinjected into the tracheal wall. Analysis of five successfully infected animals showed that 14% of PRV-GFP labeled neurons express AVP traits and 18% of transneuronally-labeled neurons contain OXT. Furthermore, the identified AVPNs innervating extrathoracic trachea receive axon terminals of the PVN neurons. The results indicate that AVP- and OXT-producing PVN cells, via direct projections to the AVPNs, could modulate cholinergic outflow to the airways, as a part of overall changes in response to stress.     

An excitatory action of 5-Hydroxytryptamine on sympathetic preganglionic neurones

Summary Preganglionic neurones in the fourth thoracic segment of anaesthetized cats fired spontaneously at a rate of about 2/sec. The effects of micro-electrophoretic administration of 5-hydroxytryptamine, noradrenaline, acetylcholine and DL-homocysteic acid on this spontaneous firing were determined. Many cells were excited by 5-hydroxytryptamine (5HT) and DL-homocysteic acid (DLH), a few were depressed by noradrenaline (NAdr), but acetylcholine (ACh) was inactive. The data are consistent with the view that 5HT and NAdr may function as excitatory and inhibitory transmitters which are released from the terminals of descending pathways in the spinal cord.National Science Foundation Postdoctoral Fellow, Riker Fellow.