Neurophysiological analysis of target-related sympathetic pathways--from animal to human: similarities and differences.

The sympathetic nervous system regulates many different target tissues in the somatic and visceral domains of the body in a differentiated manner, indicating that there exist separate sympathetic pathways that are functionally defined by their target cells. Signals generated by central integration and channelled through the preganglionic neurons into the final sympathetic pathways are precisely transmitted through the para- and prevertebral ganglia and at the neuroeffector junctions to the effector cells. Neurophysiological recordings of activity in postganglionic neurons in skin and muscle nerves using microneurography in human subjects and in skin, muscle and visceral nerves, using conventional recording techniques in anaesthetized animals, clearly show that each type of sympathetic neuron exhibits a discharge pattern that is characteristic for its target cells and, therefore, its function. These findings justify labelling the neurons as muscle vasoconstrictor, cutaneous vasoconstrictor, sudomotor, lipomotor, cardiomotor, secretomotor neurons, etc. The discharge patterns monitor aspects of the central organization of the respective sympathetic system in the neuraxis and forebrain. They can be dissected into several distinct reflexes (initiated by peripheral and central afferent inputs) and reactions connected to central signals (related to respiration, circadian and other rhythms, command signals generated in the forebrain, etc). They are functional markers for the sympathetic final pathways. These neurophysiological recordings of the discharge patterns from functionally identified neurons of sympathetic pathways in the human and in animals are the ultimate reference for all experimental investigations that aim to unravel the central organization of the sympathetic systems. The similarities of the results obtained in the in vivo studies in the human and in animals justify concluding that the principles of the central organization of sympathetic systems are similar, if not identical, at least in the neuraxis, in both species. Future progress in the analysis of the central neuronal circuits that are associated with the different final sympathetic pathways will very much depend on whether we are able to align the human models and the animal models. Human models using microneurography have the advantage to work under awake conditions. The activity in the postganglionic neurons can be correlated with various other (afferent, centrally generated) signals, effector responses, perceptions, central changes monitored by imaging methods, etc. However, human models have considerable limitations. Animal models can be divided into in vivo models and various types of reduced in vitro models. Animal models allow using various methodological approaches (e.g., neurophysiological, pharmacological, modern anatomical tracing methods; behavioural animal models; transgenic animals), which cannot be used in the human. Interaction of the research performed in the human and animals will allow to design animal models that are relevant for diseases in which the sympathetic nervous systems is involved and to trace down the underlying pathophysiological mechanisms. The scientific questions to be asked are formulated on the basis of clinical observations resulting in testable hypotheses that are investigated in the in vivo human and animal models. Results obtained in the in vivo models lead to the formulation of hypotheses that are testable in reduced in vivo and particularly in vitro animal models. Microneurographic recordings from sympathetic postganglionic fibres in the human will keep its place in the analysis of the sympathetic nervous system in health and disease although only relatively few laboratories in the world will be able to keep the standards and expertise to use this approach. Experimental investigation of the organization of the sympathetic nervous system in animal models has changed dramatically in the last 15 years. The number of in vitro models and the methodological diversity have increased. In vivo experimentation on larger animals has almost disappeared and has been replaced by experimentation on rats, which became the species for practically all types of studies on the central organization of the sympathetic nervous system.

Reflexes in sympathetic vasoconstrictor neurones arising from urinary bladder afferents are not amplified early after inflammation in the anaesthetised cat.

Pathophysiological processes in the viscera can lead to pain and hyperalgesia and exaggerated motility-regulating reflexes. This may be due to sensitisation of visceral afferents (peripheral sensitisation), which has repeatedly been shown to occur as a consequence of e.g. inflammation, and/or to sensitisation of dorsal horn neurones (central sensitisation), which is less well documented in the visceral domain. As an indicator of peripheral sensitisation, we previously analysed the responses of sacral spinal afferents after inflammation of the urinary bladder. Here, we studied reflexes in sympathetic vasoconstrictor neurones supplying skeletal muscle and skin elicited by bladder distension stimuli (vesico-sympathetic reflexes) before and after induction of bladder inflammation. Our aim was to test whether these vesico-sympathetic reflexes are amplified after inflammation in a way that would support a major functional role for post-inflammatory central sensitisation processes. Bladder inflammation was induced in anaesthetised cats by instillation of turpentine or mustard oil and vesico-sympathetic reflexes were studied 1 and 2 h after induction of the inflammation. Inflammation enhanced on-going activity in vasoconstrictor neurones supplying skeletal muscle (after 1 h to 187.6+/-36.8%, mean+/-SEM, P<0.01, and after 2 h to 139.1+/-12.9%, P<0.05, of baseline activity) and decreased it in most sympathetic neurones supplying skin (to 91.7+/-12.5%, P>0.05, and to 71.6+/-11.3%, P<0.05, respectively, of baseline activity). Relative to the altered baseline activity vesico-sympathetic reflexes to graded distension of the inflamed bladder were quantitatively unchanged with a tendency to be diminished. Thus, the changes in on-going sympathetic vasoconstrictor activity and the distension-evoked reflexes directly mirrored the afferent input from the inflamed urinary bladder into the spinal cord, i.e. no increase of the gain of these reflexes was observed. These results suggest that in the first 2 h of inflammation, peripheral sensitisation processes play the main role for hyperalgesia and hyperreflexia of the urinary bladder. In contrast, central sensitisation appears to be of little importance during this time period.

[Physiology and pathophysiology of visceral pain]. / Physiologie und Pathophysiologie viszeraler Schmerzen.

Visceral pain is diffusely localized, referred to deep somatic tissues, skin and viscera, frequently not correlated with an actual trauma, commonly correlated with strong negative affective reactions and accompanied by strong protective autonomic and motor reactions. It is correlated with the excitation of spinal (thoraco-lumbar, sacral) visceral afferents and (with a few exceptions) not with the excitation of vagal afferents. Spinal visceral afferents are polymodal and can be excited by physical and chemical stimuli. All groups of visceral afferents can be sensitized (e.g.by inflammation). Normally silent (mechanically insensitive) visceral afferents are recruited by inflammation. Individual visceral afferent neurons project in laminae I and V of the dorsal horn over several segments, medio-lateral over the entire width of the dorsal horn and to the contralateral side. Their activity is synaptically transmitted, in these and deeper laminae, to viscero-somatic convergent neurons which receive additional afferent synaptic input from skin and deep somatic tissues of the corresponding dermatomes,myotomes and sclerotomes. The mechanism of sensitization of viscerosomatic convergent neurons (central sensitization) during sensitization of spinal visceral afferents is unclear.Viscero-somatic tract neurons project to lower and upper brain stem,hypothalamus and via the thalamus to various cortex areas. Visceral nociception and pain is presumably (together with other visceral sensations and homeostatic regulations of autonomic body functions) primarily represented in the insula in the context of interoception. The insula obtains its main peripheral afferent input from lamina I neurons via the Nucleus ventromedialis posterior of the thalamus. The transmission of visceral impulses in the spinal cord is modulated by the endogenous control systems in the brain stem which are in turn under the control of cortex and limbic system.

Reflex patterns in preganglionic sympathetic neurons projecting to the superior cervical ganglion in the rat.

Reflex patterns in preganglionic neurons projecting in the cervical sympathetic trunk (CST) were analyzed in response to stimulation of various afferent systems. We focused on the question whether these preganglionic neurons can be classified into functionally distinct subpopulations. Reflex responses were elicited by stimulation of trigeminal and spinal nociceptive, thermoreceptive as well as baroreceptor and chemoreceptor afferents. Multi- and single fiber preparations were studied in baroreceptor intact and sino-aortically denervated animals. Spontaneous activity of 36 preganglionic single neurons ranged from 0.2 to 3.5 imp/s (median= 1.11 imp/s). The degree of cardiac rhythmicity (CR) in the activity of sympathetic neurons was 69.5+/-13% (mean+/-S.D.; N=52; range=39-95%). Noxious stimulation of acral skin activated the majority (67%) of sympathetic preparations by 37+/-25% (N=35) above pre-stimulus activity; 15% were inhibited. In these neurons the response to noxious stimulation of acral skin was significantly correlated with the degree of CR (P<0.001, N=52) in that neurons showing the strongest excitation to noxious stimulation displayed the strongest CR. Noxious mechanical stimulation of body trunk skin (N=60) inhibited the majority (80%) of fiber preparations tested (by 34+/-18% of pre-stimulus activity, N=48); an activation was not observed. Cold stimulation of acral (N=9) and body trunk skin (N=42) activated most fiber preparations. Trigeminal stimulation evoked a uniform reflex activation of preganglionic neurons (+79+/-73% of pre-stimulus activity, N=32). Chemoreceptor stimulation by systemic hypercapnia elicited inhibitory (-31+/-19%, N=8) as well as excitatory (+59+/-5%, N=4) responses. These results show that preganglionic sympathetic neurons projecting to target organs in the head exhibit distinct reflex patterns to stimulation of various afferent systems; however, a clear classification into different functional subgroups did not emerge. Furthermore, reflex patterns showed a segmental organization to noxious cutaneous stimulation of acral parts and body trunk reflecting a differential central integration of spinal afferent input. Compared with the cat the reflex organization of sympathetic neurons projecting to the head seems to be less differentiated in the anesthetized rat.

Respiratory rhythmicity in the activity of postganglionic neurones supplying the rat tail during hyperthermia.

It has been suggested that thermoregulatory stimulation changes respiration-related rhythmicity in the activity of postganglionic sympathetic neurones supplying the rat tail to a distinct modulation independent of respiration. To study this possibility, single and few fibre recordings were made from ten filaments split from the ventral collector nerves of the rat during whole body warming. Sympathetic activity was analysed by autocorrelation and phrenic-triggered summation. All neurones except one were gradually inhibited and lost their on-going activity above a core temperature of 39-39.5 degrees C while the frequency of the phrenic bursts increased significantly. During hyperthermia, all neurones tested exhibited a prominent respiratory modulation in their activity which, compared to normothermia, was significantly increased in strength, or even newly acquired. No other rhythm emerged. These results speak against the hypothesis that in the rat sympathetic pathways controlling the tail vasculature and thus involved in thermoregulation, during hyperthermia become controlled by central oscillators distinct from the respiratory rhythm generator. Rather, respiratory modulation appears to remain the dominant rhythm as is common for sympathetic neurones supplying other cardiovascular targets.

Sympathetic-sensory coupling after L5 spinal nerve lesion in the rat and its relation to changes in dorsal root ganglion blood flow.

Transection of the L5 spinal nerve in rats results in allodynia- and hyperalgesia-like behavior to mechanical stimulation which are thought to be mediated by ectopic activity arising in lesioned afferent neurons mainly in the dorsal root ganglion (DRG). It has been suggested that the neuropathic pain behavior is dependent on the sympathetic nervous system. In rats 3-56 days after L5 spinal nerve lesion, we tested responses of axotomized afferent fibers recorded in the dorsal root of the lesioned segment to norepinephrine (NE, 0.5 microg/kg) injected intravenously and to selective electrical stimulation of the lumbar sympathetic trunk (LST). In some experiments we measured blood flow in the DRG by laser Doppler flowmetry. The majority of lesioned afferent fibers with spontaneous activity responded to neither LST stimulation (82.4%) nor NE (71.4%). In those which did react to LST stimulation, responses occurred only at high stimulation frequencies (likely to be above the physiological range), and they could be mimicked by non-adrenergic vasoconstrictor drugs (angiotensin II, vasopressin). Excitatory responses to LST stimulation were closely correlated with the stimulation-induced phasic vasoconstrictions in the DRG. We therefore hypothesized that the activation of lesioned afferents might be brought about indirectly by an impaired blood supply to the DRG. To test this hypothesis we induced a strong and sustained baseline vasoconstriction in the DRG by blocking endothelial nitric oxide synthesis with N(G)-nitro-L-arginine methyl ester (L-NAME) applied systemically. L-NAME enhanced baseline vascular resistance in the DRG about threefold and also increased stimulation-induced vasoconstrictions. After L-NAME, the majority of axotomized neurons with spontaneous activity were activated by LST stimulation (76%) or NE (75%). Again, activations closely followed stimulation-induced phasic vasoconstrictions in the DRG provided that a critical level of vasoconstriction was exceeded. In the present study, inhibitory responses to LST stimulation were generally rare and could be reversed to activation by prolonged stimulation or after L-NAME. These results show that sympathetic-sensory coupling occurs only in a minority of axotomized afferents after L5 spinal nerve injury. Like previous studies, they cast doubt on the notion that the L5 spinal nerve lesion is a good model for sympathetically maintained pain. Since responses of lesioned afferent neurons to LST stimulation and NE could be provoked with high reliability after inducing vasoconstriction in the DRG, and since they mirrored stimulation-induced vasoconstrictions in the DRG, it appears that in this model the association of sympathetic activity with afferent discharge occurs mainly when perfusion of the DRG is impaired.

Specificity in the organization of the autonomic nervous system: a basis for precise neural regulation of homeostatic and protective body functions.

Experimental investigations of the lumbar sympathetic outflow to skin, skeletal muscle and viscera and the thoracic sympathetic outflow to the head and neck have shown that each target organ and tissue is supplied by one or two separate pathways which consists of sets of pre- and postganglionic neurons with distinct patterns of reflex activity. This probably applies to all sympathetic and parasympathetic systems. The specificity of the messages that these peripheral pathways transmit from the central nervous system arises from integration within precisely organized pathways in the neuraxis. The messages in these discrete functional pathways are transmitted to the target tissues often via organized neuroeffector junctions. Modulation in the periphery can occur within each pathway, both in ganglia and at the level of the effector organs. This organization is the basis not only for precise neural regulations of all homeostatic body functions in which the autonomic nervous system is involved but also the basis of one main component in the regulation of protective body functions: (a) Elementary defense behaviors which are organized in the mesencephalon (confrontational defense, flight, quiescence), (b) regulation of the immune system by the sympathetic nervous system, and (c) adaptive autonomic motor responses during basic emotions require precisely working autonomic, in particular sympathetic, systems. In this sense, the concept of the functioning of the sympathetic nervous system in an "all-or-none" fashion, without distinction between different effector organs, and of simple functional antagonistic organization between sympathetic and parasympathetic nervous system is misleading, inadequate and untenable.

Spontaneous activity of axotomized afferent neurons after L5 spinal nerve injury in rats.

After mechanical injury of a peripheral nerve some axotomized afferent neurons develop spontaneous activity, which is thought to trigger abnormal pain behavior in rats and neuropathic pain in humans. Here, we analysed the ectopic activity in axotomized afferent fibers recorded from the L5 dorsal root in different time periods after L5 spinal nerve lesion and the effects of sympathectomy on it. The following results were obtained: (1) Up to 6 hours after spinal nerve transection there was almost no spontaneous activity in axotomized afferents, except short-lasting injury discharges at the time of transection; (2) Three to 8 days following spinal nerve lesion, the rate of spontaneous activity was 7.3+/-7.7 imp/s (mean+/-SD, median 5.0 imp/s, n=204); 41.6% of the spontaneously active afferent neurons exhibited a bursting pattern with interspike intervals of 32.4+/-18.3 ms; (3) Twenty to 53 days after nerve lesion the rate of spontaneous activity had decreased significantly to 3.4+/-4.3 imp/s (median 2.6 imp/s, n=120). The frequency of bursting and non-bursting neurons remained roughly the same; (4) In sympathectomized rats, 15-45 days following spinal nerve lesion, the mean discharge rate was 3.8+/-4.3 imp/s (median 2. 3 imp/s, n=255). However, the percentage of bursting neurons and the intraburst frequency decreased significantly; (5) Spontaneous activity occurred in afferent A-fibers but not in afferent C-fibers. These results suggest that ectopic activity in axotomized afferent neurons develops within the first days after L5 spinal nerve lesion, decreases with time and is only marginally dependent on the sympathetic innervation. There was a positive correlation between this ectopic activity and the allodynia-like behavior in spinal nerve-lesioned rats.

Hypoventilation recruits preganglionic sympathetic fibers with inspiration-related activity in the superior cervical trunk of the rat.

Activity in preganglionic sympathetic neurons projecting in the cervical sympathetic trunk (CST) of rats was analysed with respect to changes in the pattern of the respiratory modulation during a long lasting hypoventilation. Under normal acid-base status (pH: 7.36+/-0.04, pCO2: 42.1+/-6.1 mm Hg, pO2: 135.8+/-43 mm Hg) a maximum of activity during expiration (expiration-related activity) was observed in all nerve recordings (n = 27). No other pattern of respiratory modulation was observed under this condition. Under a hypoventilation a dissociation between the duration of phrenic nerve activity and that of the inspiratory inhibition in neurons with expiration-related activity was observed as the inhibition was significantly prolonged by 49+/-24.9% and outlasted inspiration in 5/7 multifibers. When acid-base status was systematically changed (pH: 7.15+/-0.05, pCO2: 80.4+/-11.8 mm Hg, pO2: 62.8+/-17.5 mm Hg [n = 7]) by a hypoventilation lasting for several hours activity with a maximum peak during central inspiration (inspiration-related activity) emerged and disappeared when control conditions were reestablished. Neurons with expiration-related activity showed a cardiac rhythmicity (CR) of 62.5+/-14.6% (n = 27) and were inhibited to baroreceptor stimulation whereas neurons with inspiration-related activity showed no discernible CR (23.1+/-5.1%; n = 7) and were not inhibited to baroreceptor stimulation. Furthermore, expiration-related neurons were inhibited by 32.5+/-18.3% (n = 27) during noxious cutaneous stimulation while neurons with inspiration-related activity were activated by 21.5+/-12.1% (n = 7). These findings suggest that the respiratory modulation of preganglionic sympathetic activity in the CST consists of expiration-related activity in normal acid-base status. During hypoventilation neurons with inspiration-related activity are recruited. These neurons show reflex patterns distinct from expiration-related neurons and probably constitute a subgroup of sympathetic neurons which is activated under increased respiratory drive.

Frequency response characteristics of sympathetic transmission to skin vascular smooth muscles in rats.

Sympathetic modulation of cutaneous vasomotor waves in humans is most effective at frequencies up to 0.1 Hz. In contrast, sympathetic modulation of mesenteric vasomotor waves in rats is strongest in the frequency band between 0.2 and 0.75 Hz. Therefore, we addressed the question as to whether these different frequency response characteristics are due to species- or organ-specific disparities. Eleven Sprague-Dawley rats were instrumented with catheters in the carotid artery and in the jugular vein, together with electrodes on the centrally sectioned left lumbar sympathetic trunk (LST) and laser Doppler flow probes directed to the plantar surface of the skin of the left and right hind paws. In anesthetized rats, the LST was electrically stimulated at eight different stimulation frequencies, and the responses in laser Doppler blood flow were recorded in the skin of the ipsilateral and contralateral paw. At stimulation frequencies <0.2 Hz, LST stimulation induced corresponding oscillations in skin blood flow in the ipsilateral, but not in the contralateral, paw. These dynamic responses to LST stimulation in the ipsilateral paw were strongest at 0.05 and 0.075 Hz. At higher stimulation frequencies a tonic vasoconstriction was observed. It is concluded that organ-specific disparities exist in sympathetic transmission to vascular smooth muscles, whereas no species-specific differences are apparent in sympathetic transmission to cutaneous blood vessels of humans and rats.

Effects of nociceptin and nocistatin on antidromic vasodilatation in hairless skin of the rat hindlimb in vivo.

1. We tested whether nociceptin (NCE), the endogenous ligand of the opioid receptor-like 1 (ORL1) receptor, and nocistatin (NST), which reverses central NCE effects when applied intrathecally (i.t.), affect small-diameter afferent fibre-mediated vasodilatation in rat hairless skin. 2. Female Wistar rats were vagotomized. Ongoing sympathetic vasoconstrictor activity was abolished by bilateral section of the lumbar sympathetic trunk between ganglia L2 and L3. Sensory axons were selectively stimulated in the dorsal root L5 by 20 electrical impulses supramaximal for activating C-fibres at 1 Hz. Blood flow was measured on the plantar skin of the left hind paw in the L5 dermatome using laser Doppler flowmetry. 3. NCE injected intravenously (i.v.) as single boluses (1, 10 and 100 nmol kg(-1) 7 - 8 min before dorsal root stimulation (n=6) dose-dependently decreased blood pressure and local vascular resistance and suppressed antidromic vasodilatation maximally by 47% (P<0.01). When NCE was injected 2 min before stimulation (n=3), antidromic vasodilatation was reduced by 64% after NCE (1 nmol kg-1) and totally, or almost totally, abolished after the two higher doses. 4. NST (1 - 100 nmol kg(-1) i.v., n=6) was without significant effect on blood pressure and cutaneous vascular resistance. Applied 5 (n=6) or 2 min (n=3) before stimulation it also did not affect antidromic vasodilatation. NST (100 nmol kg(-1) i.v.) applied shortly before an equimolar dose of NCE did not antagonize NCE effects on vascular resistance, blood pressure and antidromic vasodilatation (n=4). 5. In conclusion, NCE inhibits antidromic vasodilatation, a component of neurogenic inflammation, in rat skin while NST is without effect. NST, at the small-diameter sensory ending, is not an effective antagonist of NCE.

Involvement of neurokinins in antidromic vasodilatation in hairy and hairless skin of the rat hindlimb.

By intravenous application of the specific neurokininl receptor antagonist SR 140333 and the specific calcitonin gene-related peptide receptor antagonist CGRP8-37 we tested to what extent neurokinins (substance P, neurokinin A) and calcitonin gene-related peptide are involved in mediating antidromic vasodilatation in skin of anaesthetized Wistar rats. The lumbar sympathetic chain was sectioned bilaterally between ganglia L2 and L3 to remove ongoing vasoconstrictor activity to the hindquarter. The left dorsal root L5 was stimulated electrically at 1 Hz with 20 pulses supramaximal for activating C-fibres to evoke antidromic vasodilatation which was measured with laser Doppler flowmetry on the glabrous plantar skin and the hairy skin of the lower hindlimb within the left L5 territory. Stimulation-induced vasodilatation was tested after applying SR 140333 (0.1 mg/kg) and CGRP8-37 (0.3 mg/kg) alone or in combination. SR 140333 delayed the onset of the vasodilatation, but did not change its amplitude. CGRP8-37 reduced the amplitude and duration of the vasodilatation, but did not affect the latency of its onset. In combination, SR 140333 potentiated the effect of CGRP8-37 on the amplitude of the vasodilatation in glabrous but not in hairy skin and CGRP8-37 potentiated the delayed onset produced by SR 140333 in both cutaneous tissues. Antidromic vasodilatation in glabrous skin was almost totally blocked by SR 140333 (0.1 mg/kg) in combination with CGRP8-37 (0.45 mg/kg), but a substantial dilatation remained in hairy skin. It is concluded that in rat glabrous skin the vasodilatation evoked by a low level of activity in small-diameter primary afferents is likely to result from the release and synergistic action of neurokinins (substance P and/or neurokinin A) and calcitonin gene-related peptide, while in hairy skin neurokinins are involved to a minor extent only.

Rhythmicity in single fiber postganglionic activity supplying the rat tail.

Rhythmicity in single fiber postganglionic activity supplying the rat tail. The temporal pattern of ongoing sympathetic vasoconstrictor activity may play an important role for neurovascular transmission. Here we analyzed the activity of postganglionic fibers projecting into the ventral collector nerve of anesthetized and artificially ventilated vagotomized Wistar rats with respect to the presence of rhythmic firing under normocapnic conditions. Most of the fibers studied were likely vasoconstrictor and involved in thermoregulation. Accumulated histograms of sympathetic activity were produced synchronized with the electrocardiogram to detect cardiac rhythmicity, with phrenic nerve activity to detect modulation with the central respiratory cycle, and with tracheal pressure to uncover a reflex modulation associated with artificial ventilation. Sympathetic activity, phrenic activity, and tracheal pressure also were examined by spectral analysis and autocorrelation to detect rhythmicities distinct from respiration. Twenty-seven filaments containing two to seven fibers with spontaneous activity and 51 single fibers were analyzed. Ongoing activity was 1.12 +/- 0.65 imp/s (mean +/- SD, n = 51); conduction velocity was 0.62 +/- 0.06 m/s (n = 30). Cardiac rhythmicity in sympathetic activity was weak (46.2 +/- 16.4%). The dominant rhythm in the activity of 19/27 few-fiber preparations and 37/51 single fibers corresponded to the central respiratory cycle. The pattern consisted of an inhibition during inspiration and an activation in expiration. In 10/19 few-fiber preparations and 21/37 single fibers of this group, there was also a concomitant, less prominent rhythm related to artificial ventilation. By contrast, 8/27 few-fiber preparations and 11/51 single fibers exhibited a dominant pump-related modulation, whereas phrenic-related rhythmicity was subordinate. The dominant rhythm in the activity of two single fibers was related to neither central respiration nor artificial ventilation. We conclude that the ongoing activity of most postganglionic neurons supplying the rat tail is modulated by the central respiratory rhythm generator, suggesting that changes in respiratory drive may alter perfusion of the tail and therefore heat dissipation. Reflex modulation in parallel with artificial ventilation, independent of vagal afferents and possibly due to ventilatory changes of baroreceptor activity, is also an important source of rhythmicity in these neurons.

Synaptic responses evoked by lower urinary tract stimulation in superior cervical ganglion cells in the rat.

PURPOSE: Stimulation of afferent neurons from the urinary tract can evoke powerful cardiovascular reflexes in animals and in humans. Here we tested whether and to what extent postganglionic sympathetic neurons projecting to the head and neck are involved in these reflexes. MATERIALS AND METHODS: In adult anesthetized female Wistar rats, reflex changes in synaptic activity elicited from the lower urinary tract were recorded in neurons of the superior cervical ganglion using intracellular recording techniques. RESULTS: Gentle movements of the urethral cannula and/or slow injections of 0.3 to 0.6 ml. saline into the bladder modified synaptic activity in 75% of neurons tested: 11/15 neurons tested showed reflexes to urethral stimulation (8 excitatory, 3 inhibitory) and 8/12 neurons responded to bladder distension (5 excitatory, 3 inhibitory). Five neurons showed a reciprocal response pattern to the two stimuli. Suprathreshold and subthreshold inputs to a given cell mostly showed the same type of response. There was no evidence that urinary tract stimulation recruited preganglionic inputs that did not have ongoing activity prior to the stimulus. CONCLUSIONS: Reflexes from the lower urinary tract clearly reached sympathetic neurons located in remote segments. The response incidence in the population studied suggests that most sympathetic neurons involved in cardiovascular regulation participate in these reflexes. The different reflex patterns probably occur in neurons with different functional targets in the head and neck.

Changes in the action potential in sensory neurones after peripheral axotomy in vivo.

Following nerve injury, modified somatic ion channels may underlie ectopic activity in axotomized A-type neurones in dorsal root ganglia (DRGs) leading to abnormal pain signalling. Using intracellular microelectrodes both in vivo and in vitro, action potentials (APs) were recorded in rat DRG neurones classified by axonal conduction velocity. After lesions to L5 spinal or sciatic nerves, APs in both A alpha/beta and A delta cells were wider, and those in A alpha/beta neurones more frequently showed inflections during repolarization, than APs in cells in undamaged ganglia. AP amplitudes and dV/dt(max) were not significantly altered by axotomy. These results confirm previous observations in intact ganglia in vitro but differ from those reported for dissociated neurones using patch recording techniques.

Lumbar sympathectomy failed to reverse mechanical allodynia- and hyperalgesia-like behavior in rats with L5 spinal nerve injury.

The L5 spinal nerve ligation model of neuropathic pain in rats has been proposed as a model for sympathetically maintained pain (SMP) based on the effects of surgical or chemical sympathectomy on nerve injury induced behavior. In an attempt to confirm that the lesion produces an animal model of SMP, surgical sympathectomies were independently conducted in two different laboratories (Johns Hopkins and University Kiel) using male Sprague-Dawley (n = 30) or Wistar rats (n = 14). The L5 spinal nerve was ligated or cut and ligated. Using von Frey hairs, paw withdrawal threshold and incidence of paw withdrawal were tested concurrently before and after the sympathectomy. The sympathectomy was either verified by (a) glyoxylic acid staining of peripheral blood vessels of the hindpaw or (b) skin temperature measurements of the hindpaws. To blind the experimenter, surgeries and behavioral tests were performed by two different investigators and a sham sympathectomy was performed at Johns Hopkins. Decreased paw withdrawal thresholds and increased frequencies of paw withdrawal on the lesioned side were observed after the L5 lesion. Thus, the L5 spinal nerve ligation resulted in behavioral signs of allodynia and hyperalgesia to mechanical stimuli. Lumbar surgical sympathectomy 1-3 weeks after the lesion or prior to lesion with bilateral removal of the sympathetic ganglia L2-L4, however, did not reverse or prevent the behavioral changes induced by the nerve injury. The lack of effect of the sympathectomies was independent of the testing paradigm used. Experiments in Wistar and Sprague-Dawley rats yielded the same results. Potential reasons for the discrepancies between the present study and earlier reports are discussed. These results indicate that an L5 spinal nerve injury rat model is not a reliable model for SMP.

Responses of axotomized afferents to blockade of nitric oxide synthesis after spinal nerve lesion in the rat.

Lesioned afferents were tested for their responses to blockade of nitric oxide synthesis in the spinal nerve L5 lesion model for neuropathic pain in Wistar rats. Seven single fibers with spontaneous activity split from dorsal root L5 showed no response after non-selective blockade of nitric oxide synthesis with N(G)-nitro-L-arginine methyl ester whereas five were excited after 5-7 min. Three previously silent units were recruited. Blood flow in the dorsal root ganglion decreased. None of fifteen axotomized afferents tested responded to selective blockade of neuronal nitric oxide synthesis with 7-nitroindazole. It is concluded that neuronal nitric oxide is not involved in the generation of spontaneous activity in axotomized afferent neurons in this model. We suggest that the vasoconstriction induced by blockade of endothelial nitric oxide may be responsible for the excitatory responses.

Analysis of the periodicity of synaptic events in neurones in the superior cervical ganglion of anaesthetized rats.

1. The patterns of on-going synaptic events recorded intracellularly in neurones of superior cervical ganglia (SCG)of anaesthetized female rats were analysed by constructing inter-event interval histograms, autocorrelograms, ln-survivor curves and histograms triggered by the arterial pulse wave and by the intercostal EMG. 2. In 11/12 cells with on-going frequencies > 0.5 Hz, one or two inputs were strong (i.e. always suprathreshold). In five cells, action potentials also arose from synaptic potentials with amplitudes close to threshold. 3. Synaptic events in 5/11 neurones tested were phase-related to the arterial pressure wave (i.e. had cardiac rhythmicity, CR). 4. Synaptic events in 9/10 neurones tested (including all with CR) were phase-related to the intercostal EMG and/or their autocorrelograms showed peaks at multiples of the respiratory interval (i.e. had respiratory rhythmicity, RR). 5. The intervals between all synaptic events were exponentially distributed in 8/12 neurones although intervals between single strong events showed peaks related to the respiratory cycle. Bursts occurred only by chance. 6. Event patterns could be simulated by combining events from several respiration-modulated inputs with their timing distributed over nearly half the cycle. From the simulations, the mean number of active preganglionic inputs was estimated to be approximately 6 with mean discharge frequency approximately 0.4 Hz. 7. We conclude that, in the spontaneously breathing anaesthetized rat, most preganglionic neurones to the SCG fire with relatively low probability in relation to the respiratory cycle. Rhythms in a postganglionic neurone reflect the activity of its suprathreshold preganglionic inputs.

Functional evidence for the differential control of superficial and deep blood vessels by sympathetic vasoconstrictor and primary afferent vasodilator fibres in rat hairless skin.

We quantitatively investigated sympathetic vasoconstriction and antidromic vasodilation mediated by small-diameter primary afferents on the plantar hairless skin of the hindpaws in Wistar rats using laser Doppler (LD) flowmetry and an infrared thermometer. Sympathetic vasoconstriction was elicited by electrical stimulation of the centrally cut ipsilateral lumbar sympathetic trunk (LST) with 50-s trains at 0.1-20 Hz. Antidromic vasodilation was evoked by electrical stimulation of the dorsal root (DR) L5 with 20-s or 50-s trains at 1-4 Hz. Cutting the LST resulted in increases in skin temperature (SKT) by 6.1 +/- 1.0 degrees C (mean +/- SEM) and in LD flow by 128 +/- 20%. Stimulation of the LST resulted in a graded decrease in LD flow and SKT that was most pronounced between 0 and 0.1 Hz. However, DR stimulation evoked a large increase in LD flow but only little change in SKT in rats with sectioned LST. When the DR was stimulated either in animals with intact LST or during continuous stimulation of vasoconstrictor fibres in the sectioned LST, i.e. while baseline temperature was relatively low (26.3 +/- 1.1 degrees C), DR stimulation still resulted in large increases in LD flow, but only minor changes in SKT. These results suggest that blood flow through both deep and superficial layers of rat hairless skin is regulated by activity in sympathetic postganglionic vasoconstrictor fibres, whereas small-diameter primary afferent fibres appear to influence predominantly the blood flow through superficial layers of rat plantar skin.