Parabrachial nucleus influences the control of normal urinary bladder function and the response to bladder irritation in rats.

The contribution of the parabrachial nucleus to the mediation of bladder contraction was examined in the rat. Constant infusion (0.1 ml/min) of saline or 0.2% acetic acid evoked normal or abnormal bladder contractions, respectively. Single unit activity was recorded in the parabrachial nucleus with tungsten microelectrodes. Seven units with activity that was correlated with bladder contraction during saline infusion were located in the lateral subnuclei and three units were located in the medial subnuclei of the parabrachial nucleus. Twelve units with activity that was correlated with abnormal bladder contractions were found widely distributed in the parabrachial nucleus. An inverse correlation of activity to normal or abnormal bladder contractions was identified in 11 units in the parabrachial nucleus. Pressure injection of 5 mM CoCl(2) into the parabrachial nucleus was used to block synaptic transmission unilaterally. Normal bladder contractions evoked by saline infusion were disrupted by 5 of 10 injections, 4 of them in the medial subnuclei of the parabrachial nucleus and one in the lateral subnuclei. Abnormal bladder contractions were converted to a normal pattern in nine experiments where CoCl(2) injections lay in the lateral subnuclei of the parabrachial nucleus. In five experiments, CoCl(2) disrupted abnormal bladder contractions; four effective sites were located in the lateral subnucleus and one lay in the medial subnucleus of the parabrachial nucleus. These data demonstrated that single units responding to both normal and abnormal contractions were located throughout the parabrachial nuclei whereas the lateral subnuclei play a predominant role in mediation of abnormal bladder contractions and the medial subnuclei play a predominant role in the mediation of normal bladder contractions.

Chronic administration of amitriptyline and caffeine in a rat model of neuropathic pain: multiple interactions.

This study was designed to determine (1) whether chronic amitriptyline administration was effective in alleviating symptoms of neuropathic pain in a rat model of spinal nerve injury, and (2) whether the effect of amitriptyline involved manipulation of endogenous adenosine, by determining the effect of caffeine, a non-selective adenosine A(1) and A(2) receptor antagonist, on its actions. Nerve injury was produced by unilateral spinal nerve ligation of the fifth and sixth lumbar nerves distal to the dorsal root ganglion, and this resulted in stimulus-evoked thermal hyperalgesia and static tactile mechanical allodynia. Animals received pre- and post-surgical intraperitoneal doses of amitriptyline (10 mg/kg) and caffeine (7.5 mg/kg), alone or in combination, and following surgery, were administered amitriptyline (15-18 mg/kg/day) and caffeine (6-8 mg/kg/day), alone or in combination, in the drinking water. Rats were tested for thermal reaction latencies and static tactile thresholds at 7, 14 and 21 days following surgery. In the paw ipsilateral to the nerve ligation, chronic amitriptyline administration consistently decreased the thermal hyperalgesia produced by spinal nerve ligation over a 3-week period, and this effect was blocked by concomitant caffeine administration at all time intervals. In the contralateral paw, thermal withdrawal latencies were more variable, with the most reproducible finding being a reduction in thermal thresholds in the amitriptyline-caffeine combination group. There was no effect by either drug or the drug combination on the static tactile allodynia produced by spinal nerve ligation in the ipsilateral paw. However, chronic amitriptyline administration induced a tactile hyperaesthesia in the contralateral paw at all time intervals, and this effect was exacerbated by concomitant chronic caffeine administration. The results of this study indicate that chronic administration of amitriptyline is effective in alleviating thermal hyperalgesia, but not static tactile allodynia, in the hindpaw ipsilateral to nerve injury, and the block of this effect by caffeine suggests that this effect is partially achieved through manipulation of endogenous adenosine systems. Additionally, chronic amitriptyline administration induces contralateral hyperaesthetic responses that are augmented by caffeine. Both the symptom-specific effect, and adenosine involvement in amitriptyline action may be important considerations governing its use in neuropathic pain.

Induction of heat shock proteins and motor function deficits after focal cerebellar injury.

A weight drop model of focal cerebellar injury was used to identify heat shock protein induction and motor function deficits in the anesthetized, adult male, Sprague-Dawley rat. All animals were trained on a beam walking test prior to surgery. Groups of animals received severe, mild or sham weight drop injury to the lateral/paravermal region of the cerebellum. The mild and sham-injured animals showed no motor deficits in the beam walking test, whereas animals with severe cerebellar injury showed significant motor deficits in the beam walking test that approached recovery of motor function 20 days after injury. Following severe injury, induction of heat shock protein of 27kDa was observed in Purkinje cells and in neurons of the deep cerebellar nuclei, as well as Bergmann glial cells, glial cells located in the granule cell layer and the underlying white matter. Following mild injury, heat shock protein of 27kDa induction was observed in Purkinje cells and glial cells, but not in neurons of the deep cerebellar nuclei. The labeled Purkinje cells were widely distributed in the ipsilateral cerebellar cortex. Many of the glial cells that were immunostained with heat shock protein of 27kDa co-localized with cells immunoreactive for glial fibrillary acidic protein. After severe injury, heat shock protein of 72kDa was localized mainly in granule cells at the site of the trauma and in the ipsilateral deep cerebellar nuclei whereas, after mild injury, light labeling was observed only in the granule cell layer. The results demonstrate that focal cerebellar injury has profound effects on motor behavior and induces different families of heat shock proteins in specific groups of neurons and glial cells in the cerebellum.

Micturition evoked by glutamate microinjection in the ventrolateral periaqueductal gray is mediated through Barrington's nucleus in the rat.

Neural tracing experiments have demonstrated a direct spinal projection to Barrington's nucleus and a possible indirect pathway to Barrington's nucleus via the periaqueductal gray. We sought to identify the role of the periaqueductal gray matter in micturition in urethane-anesthetized rats. Blockade of micturition by focal injection of cobalt chloride was used to identify sites critical to micturition. These sites were located near the ventral margin of the caudal ventrolateral periaqueductal gray and in Barrington's nucleus. L-Glutamate injections into caudal regions of the periaqueductal gray evoked bladder contraction with coordinated sphincter activation. Additional L-glutamate sites with a similar pattern of response and sites where sphincter activation was produced without bladder contraction were found more rostrally and dorsally in the periaqueductal gray. Activation of bladder contractions by L-glutamate injection in the ventrolateral periaqueductal gray was blocked by prior injection of cobalt chloride into Barrington's nucleus. From these data we propose that ventrolateral periaqueductal gray is functionally important to micturition in the urethane-anesthetized rat. Further, we have shown that a periaqueductal gray to Barrington's nucleus pathway is functionally relevant to central mediation of bladder contraction.

Comparative behavioural toxicity of domoic acid and kainic acid in neonatal rats.

Cumulative behavioural toxicity was measured in groups of male and female rat pups (n=6/sex) at different stages of postnatal development. Dose-response curves (DRCs) for toxicity produced by domoic acid (DOM) were generated using animals on postnatal days (PND) 0, 5, 14, and 22, using a behavioural rating scale. In a subsequent experiment, DRCs for toxicity generated by either DOM or kainic acid were produced in rats at PND 8 and 14 for comparison between the two toxins. DOM was found to be a very potent neurotoxin in newborn rats and the potency of DOM progressively decreased with increasing age (interpolated ED(50)=0.12, 0.15, 0.30, and 1.06 mg/kg at PND 0, 5, 14, and 22, respectively). In addition, the patterns of behavioural expression were found to differ with age. Comparisons between DOM and kainic acid revealed that DOM was approximately six-fold more potent than kainate at both PND 8 and PND 14 and that both toxins were approximately two-fold less potent in PND 14 rats, compared to PND 8. This implies that the mechanism(s) responsible for reduced potency is/are similar between the two compounds. Consistent with previous reports, however, there were both similarities and differences in the observed patterns of behavioural toxicity produced by the two toxins at both ages.

Conditioning effects of repetitive mild neurotrauma on motor function in an animal model of focal brain injury.

A weight drop model of brain injury was used to determine the effects of repetitive mild brain injury on motor function, heat shock protein and glial fibrillary acidic protein expression in the anesthetized, adult male, Sprague-Dawley rat. Repetitive mild brain injury was produced when animals received a series of three mild injuries spaced three days apart. A separate group of repetitive mild injured animals also received a subsequent severe brain injury between three and five days after the last mild injury. All animals were trained on a beam-walking test prior to surgery. The mild, repetitive mild and repetitive mild plus severe brain injury groups showed no motor deficits in the beam-walking test, whereas the animals with only severe brain injury showed significant motor deficits (increase in number of footslips) in the beam-walking test that recovered within eight days after injury. Both repetitive mild plus severe injury and severe injury only animals had cortical necrotic cavities of similar size in the region of the hindlimb motor cortex. Both the repetitive mild and severe brain-injured animals had marked heat shock protein 27kDa and glial fibrillary acidic protein staining in the cerebral cortex. Fluoro-Jade, heat shock protein 27kDa and 72kDa labeling indicated that there were widespread effects on cortical, subcortical and spinal neurons and glial cells after repetitive mild brain injury. These results suggest that repetitive mild brain injury conditions the brain so that subsequent brain injury at the same site has no effect on motor function. Furthermore, repetitive mild injury-induced activation of processes distant to the primary injury site may have a role in activation of secondary sites involved in recovery of motor function.

Visceral afferent activation-induced changes in sympathetic nerve activity and baroreflex sensitivity.

The following experiments were done to determine whether changes in baroreflex sensitivity evoked by cervical vagus nerve stimulation are due to sympathoexcitation mediated by the parabrachial nucleus. The relative contribution of cardiopulmonary and general gastric afferents within the cervical vagus nerve to the depression in baroreflex sensitivity are also investigated. Male Sprague-Dawley rats anesthetized with thiobutabarbital sodium (50 mg/kg) were instrumented to measure blood pressure and heart rate or for the continuous monitoring of renal sympathetic nerve activity. Baroreflex sensitivity was measured using bolus injections of phenylephrine. Electrical stimulation of the cervical vagus (with or without the aortic depressor nerve) or the abdominal vagus nerve produced a significant increase in renal nerve activity and a decrease in baroreflex sensitivity. Both of these effects were blocked after the microinjection of lidocaine into the parabrachial nucleus before nerve stimulation. Therefore, we conclude that an increase in the activity of cardiac, pulmonary, or general gastric afferents mediated the increased sympathetic output and decreased baroreflex sensitivity via a pathway involving the parabrachial nucleus.

Volume-evoked micturition reflex is mediated by the ventrolateral periaqueductal gray in anesthetized rats.

The central pathway of the micturition reflex in the rat was investigated functionally by acute blockade of synaptic neurotransmission using microinjection of cobalt chloride into the periaqueductal gray or pontine tegmental region. In 27 urethan-anesthetized (1.2 g/kg ip) rats, the bladder pressure response to continuous infusion of the bladder with saline (0.1-0.25 ml/min) was assessed. Electromyographic activity of external urethral sphincter and arterial blood pressure were also monitored. Bladder contractions and external urethral sphincter activity were reversibly attenuated after unilateral or bilateral stereotaxic injections of 10 mM cobalt chloride into the caudal (bregma -7.80 to -8.80) ventrolateral periaqueductal gray as well as into Barrington's nucleus. Blood pressure was not affected by injection into either area. The results demonstrate that the caudal ventrolateral periaqueductal gray, in addition to Barrington's nucleus, is a critical part of the long-routed micturition reflex circuitry in the anesthetized rat.

Convergent prefrontal cortex and mamillary body projections to the medial pontine nuclei: a light and electron microscopic study in the rat.

To study the convergence of medial prefrontal cortex and mamillary body projections to the medial pontine nuclei, light and electron microscopic, neuroanatomical, tract-tracing experiments were performed. Injections of horseradish peroxidase conjugated to wheat germ agglutinin (WGA-HRP), biotin conjugated to dextran (BD), or rhodamine conjugated to dextran (RD) were made individually or in combinations into the cerebral cortex, hypothalamus, or pons. In addition, injections of WGA-HRP into the medial prefrontal cortex and electrolytic lesions of the mamillary body were made to study the synaptology of afferent projections to the pontine nuclei. In the light microscopic studies, injections of WGA-HRP into the rostromedial pontine nuclei produced dense, retrograde labeling both in the dorsal peduncular area of the medial prefrontal cortex and in the medial mamillary nucleus, pars medialis. Injections of the anterograde tracers BD and RD into the medial prefrontal cortex and the medial mamillary nuclei, respectively, resulted in partially overlapping terminal fields in the rostromedial pontine nuclei. In the electron microscopic studies, injections of WGA-HRP into the dorsal peduncular area and electrolytic lesions of the mamillary body produced anterogradely labeled axon terminals and degenerating axon terminals that synapsed on the same dendrites or neuronal somata in the rostromedial pontine nuclei. The results demonstrate that the medial prefrontal cortex and the medial mamillary nuclei have partially overlapping projections to the rostromedial pontine nuclei and implicate precerebellar relay nuclei in the integration of limbic and/or autonomic functions mediated by convergent projections from the cerebral cortex and the hypothalamus.

Trigeminal-reticular connections: possible pathways for nociception-induced cardiovascular reflex responses in the rat.

Cardiovascular regulatory neurons of the ventral medulla and pons are thought to have an important role in the mediation of trigeminal nociception-induced reflex cardiovascular responses. However, the neural pathways that link the spinal trigeminal nucleus with ventral medullary and pontine autonomic cell groups are poorly understood. The present study utilized injections of the highly sensitive anterograde tracer substance biotinylated dextran combined with immunocytochemistry for tyrosine hydroxylase, the synthesizing enzyme for catecholamines, to investigate the distribution and morphology of projections from the spinal trigeminal subnucleus caudalis to ventral medullary and pontine catecholaminergic cell groups. Injection of biotylinated dextran into the dorsal subnucleus caudalis produced dense anterograde labeling in dorsal regions of the medullary and pontine reticular formation including the dorsal medullary reticular field, the parvicellular reticular field, and the parvicellular reticular field pars anterior. In the ventral medullary and pontine reticular formation, light anterograde labeling tended to be distributed in close proximity to the distal dendrites of catecholaminergic neurons located in the C1, A1, and A5 regions. Injections of anterograde tracer into the dorsal medullary reticular field produced dense anterograde labeling in the ventral medullary and pontine reticular formation. Numerous terminal-like varicosities were observed in close proximity to catecholaminergic neurons located in the C1, A1, and A5 regions. These data suggest that trigeminal pain-induced reflex cardiovascular responses involve indirect projections that terminate in the dorsal medullary and pontine reticular formation before reaching ventral medullary and pontine catecholaminergic cell groups known to be involved in cardiovascular regulation.

Trigeminal autonomic pathways involved in nociception-induced reflex cardiovascular responses.

Reflex cardiovascular responses elicited by noxious oro-facial stimulation are well known but the neural pathways that underlie trigeminal cardiovascular reflex reactions remain to be elucidated. In previous studies, we have shown that noxious electrical stimulation of the mandibular incisor in the anesthetized rat elicits increases in mean arterial blood pressure and heart rate (Allen, G.V., Barbrick, B. and Esser, M.J., Trigeminal parabrachial connections: possible pathway for nociception-induced cardiovascular reflex responses, Brain Res., 715 (1996) 125-135). In this study, microinjections of the presynaptic blocker, cobalt chloride, or the anesthetic agent, lidocaine, were made into selected brainstem sites to identify neural pathways that are involved in mediation of the reflex pressor responses. Ipsilateral and bilateral injections of chemical blocker into the dorsomedial spinal trigeminal nucleus, pars caudalis, lateral parabrachial nucleus and the rostral ventral lateral medulla/caudal A5 region attenuated the reflex cardiovascular response. Bilateral injections of cobalt chloride into the dorsomedial subnucleus caudalis resulted in 70-100% attenuation of the reflex pressor response. Bilateral injections of cobalt chloride and/or lidocaine into the lateral parabrachial nucleus or the rostral ventral lateral medulla/A5 region resulted in 43-57% and 44-100% attenuation of the reflex pressor response, respectively. There were no significant differences in the degree or duration of attenuation of the reflex pressor responses produced by cobalt chloride compared to that produced by lidocaine injections. The reflex pressor responses usually returned to baseline levels approximately 60 min following injection of the chemical blocker substance. The results indicate that noxious electrical stimulation of the mandibular incisor elicits a reflex increase in mean arterial blood pressure which is initially mediated in the dorsomedial spinal trigeminal nucleus, pars caudalis and is subsequently mediated in the lateral parabrachial nucleus and the rostral ventral lateral medulla/caudal A5 region.

Trigeminal-parabrachial connections: possible pathway for nociception-induced cardiovascular reflex responses.

Noxious stimulation of dental nerves elicits marked changes in cardiovascular function. In order to investigate central pathways mediating reflex changes in cardiovascular activity, immunohistochemical localization of cells expressing the immediate-early gene, c-fos, was used to identify central nervous responding to noxious electrical stimulation of mandibular, incisor tooth dentin or chemical (capsaicin) stimulation of tooth pulp in the anesthetized rat. Injections of Fluoro-Gold were made in the lateral parabrachial region to identify efferent projections from the spinal trigeminal nucleus. Electrical and chemical stimulation produced similar patterns of Fos-positive neurons in the spinal trigeminal nucleus: subnuclei caudalis, interpolaris and oralis. Fos-positive neurons were most dense in laminae I and II of the dorsomedial subnucleus caudalis with fewer Fos-positive neurons located in the interpolaris and oralis subnuclei. Sham stimulation of tooth dentin and control vehicle injections into the tooth pulp resulted in either a few weakly stained or no Fos-positive neurons in the spinal trigeminal nucleus. Cell bodies double labeled with Fluro-Gold following injections into the parabrachial region and Fos-protein subsequent to electrical stimulation of incisor tooth were present in all three subnuclei of the spinal trigeminal nucleus. The largest number of Fos-positive neurons with efferent projections to the lateral parabrachial region were located in subnucleus caudalis (32.2 +/- 5.3 S.E.M.) and fewer were located in the interpolaris (0.4 +/- 0.4 S.E.M.) and oralis (19.8 +/- 3.5 S.E.M.) subnuclei. The results demonstrate that nociceptive dental input received by the three subnuclei of the spinal trigeminal nucleus, particularly the subnucleus caudalis, is relayed to the lateral parabrachial nucleus.

Neurotensin in the lateral hypothalamic area: origin and function.

The origin of neurotensin in the lateral hypothalamus was investigated by means of fluorescent retrograde tract tracing and neurotensin-like immunoreactivity. Following fluorescent retrograde tract tracing with FluoroGold combined with neurotensin immunohistochemistry in the rat brain, numerous neurotensin-immunoreactive neurons with projections to the posterior lateral hypothalamic area were identified in the central nucleus of the amygdala, perifornical area and the parabrachial nucleus. Fewer numbers of neurotensin-positive neurons with projections to the lateral hypothalamic area were observed in the bed nucleus of the stria terminalis, lateral septal nucleus, medial preoptic area, peri- and paraventricular nuclei of the hypothalamus, anterior lateral hypothalamic area and dorsal raphe nucleus. In addition, the role of neurotensin in the modulation of autonomic regulatory input from the insula was investigated. The lateral hypothalamic area was surveyed for single units responding to electrical stimulation (500-900 microA, 0.5 Hz) of sites in the insular cortex from which cardiovascular pressor or depressor responses could be elicited. These units were tested for the influence of neurotensin on responses to stimulation of the insular cortex. Of 60 spontaneously firing neurons, 27 units responded to electrical stimulation of cardiovascular sites in the insula. Of the units responding to stimulation of cardiovascular sites in the insula, 14 units showed excitation only, 10 units showed excitation followed by inhibition and three units showed inhibition. Iontophoresis of 0.1-1.0 mM neurotensin (25-100 nA, pH 5.0-6.0) potentiated six of the excitatory responses and showed no effect on the inhibitory responses. In addition, nine neurons showed an increase in spontaneous activity with iontophoresis of neurotensin. Of these neurons, three were excited by insular stimulation and six did not respond. These findings indicate the likely origin of neurotensin in the lateral hypothalamic area and demonstrate that neurotensin has a role in the modulation of some of the cardiovascular regulatory input from the insular cortex.

Neurochemical changes following occlusion of the middle cerebral artery in rats.

We have developed a stroke model involving middle cerebral artery occlusion in the rat which elicits changes in cardiac and autonomic variables that are similar to those observed clinically. It is likely that these neurogenic autonomic responses are mediated by changes in neurotransmitter systems subsequent to the stroke. This possibility was investigated by examining changes in immunohistochemical staining for tyrosine hydroxylase, neuropeptide Y, leu-enkephalin, neurotoxins and dynorphin following middle cerebral artery occlusion in the rat. Computerized image analysis was used to provide semi-quantitative measurements of the changes. The ischemic region was centered primarily in the insular cortex. The results indicate that there are significant increases in immunostaining for tyrosine hydroxylase and neuropeptide Y in the insular cortex within the peri-infarct region. Neuropeptide Y staining was also significantly increased in the basolateral nucleus of the amygdala, ipsilateral to the middle cerebral artery occlusion, which did not appear to be included in the infarct. Leu-enkephalin, neurotensin and dynorphin staining was significantly elevated in the central nucleus of the amygdala ipsilateral to the occlusion of the middle cerebral artery. These neurochemical changes are discussed as possible mechanisms mediating the cardiac and autonomic consequences of stroke or as part of a process to provide neuro-protection following focal cerebral ischemia.

Monoamines in the parabrachial nucleus of the cardiomyopathic hamster.

The development of dystrophic cardiac muscle is related to increases in sympathetic nervous system activity but little is known regarding possible central neural mechanisms that may be involved in cardiomyopathy. The inbred cardiomyopathic hamster is an animal model for studying the development and mechanisms of necrosis in cardiac muscle which resemble non-vascular myocardial diseases of man. Because monoamines are known to play a major role in central regulation of the cardiovascular system, we compared the distribution and density of tyrosine hydroxylase (TH) and 5-hydroxytryptamine (5-HT) immunostaining in the brains of cardiomyopathic hamsters (strain CHF-146), a related strain (CHF-148) of non-cardiomyopathic albino hamsters, and golden Syrian hamsters for possible differences in neurochemical organization. At the time of sacrifice, the cardiomyopathic hamsters exhibit small, calcified lesions on the surface of the ventricular cardiac muscle (early necrotic phase). Brain sections from each group were processed identically and simultaneously. The results show that there were significant differences among strains in the parabrachial nucleus with respect to the two neurochemicals examined. In golden Syrian and albino hamsters, TH and 5-HT immunoreactive axons were lightly-to-moderately stained in the lateral parabrachial nucleus. In the cardiomyopathic hamster, there were significantly more densely stained TH and 5-HT immunoreactive axons in the lateral parabrachial nucleus, in particular the inner part of the external lateral subnucleus. Because the lateral parabrachial nucleus, including the external lateral subnucleus, is known to be involved in regulation of the cardiovascular system, the differential distribution of TH and 5-HT in the parabrachial nucleus of cardiomyopathic hamsters in comparison to normal hamsters suggests that the parabrachial nucleus could be involved in sympathetic mechanisms related to the development of necrosis in cardiac muscle of the cardiomyopathic hamster.

Serotoninergic and nonserotoninergic neurons in the medullary raphe system have axon collateral projections to autonomic and somatic cell groups in the medulla and spinal cord.

Fluorescent double retrograde-tracing studies combined with fluorescent immunostaining for serotonin were carried out to determine the potential patterns of divergence in axonal projections to autonomic and somatic motor sites from medullary raphe and parapyramidal neurons. Injections (20-60 nl) of combinations of fluorescent retrograde tracers (Fast Blue, fluoro-gold, green latex microspheres, Diamidino Yellow) were made into the intermediolateral cell column (IML) of the spinal cord and the brainstem lateral tegmental field or ventral horn of the lumbar spinal cord of male Wistar rats. The animals were perfused after a 7-10-day survival period, and the brains were removed, sectioned (50 microns), and immunostained for serotonin. Following injections of different retrograde-tracer substances into the IML of the thoracic cord and the ventral horn of the lumbar cord, 36% of the neurons with axon collateral projections to the IML and the lumbar ventral horn were serotoninergic. Following injections of different retrograde-tracer substances into the IML and the lateral tegmental field, 26% of the neurons with axon collateral projections to the IML and the lateral tegmental field were serotoninergic. Many of the medullary neurons with projections to the lateral tegmental field and the lumbar cord were located dorsal and lateral to those neurons with projections to the IML. The results indicate that serotoninergic and nonserotoninergic neurons of the midline raphe system and parapyramidal region have axon collateral branches to the IML and the lateral tegmental field or the IML and the lumbar ventral horn. These projection neurons may form the anatomical substrate for the integration of autonomic and somatic motor activity.

Functional and anatomical organization of cardiovascular pressor and depressor sites in the lateral hypothalamic area. II. Ascending projections.

Microinjections of L-glutamate or D,L-homocysteic acid were used to stimulate cell bodies in the region of the lateral hypothalamic area (LHA) selectively. Subsequent iontophoretic injections of Phaseolus vulgaris-leucoagglutinin or pressure injections of wheat germ agglutinin-horseradish peroxidase were made into regions containing identified pressor and depressor sites and their connections with the forebrain and cerebral cortex were traced. The results indicate that decreases in blood pressure (10-45 mm Hg) and heart rate (20-70 bpm) could be elicited from tuberal (LHAt) and posterior (LHAp) sites in the LHA and that these regions have ascending projections to the insular cortex, the ventral forebrain including the septal-diagonal band of Broca complex, the ventral palladium, substantia innominata, amygdala, and the lateral preoptic area. In contrast, increases in blood pressure (10-40 mm Hg) and heart rate (20-70 bpm) were elicited primarily from neurons located adjacent to the fornix in the perifornical area (PFA). Injections of tract tracers into this region produced terminal labeling that differed markedly from the pattern seen following injections of tracer into depressor sites in the LHA. In addition, the pattern of anterograde labeling seen following injections of tracer into the anterior PFA differed from that seen following injections of tracer into the posterior PFA. Injections of tracer into the anterior PFA resulted in dense terminal labeling in the medial preoptic area and the parvicellular paraventricular nucleus of the hypothalamus whereas injections into the posterior PFA resulted in dense terminal labeling in the lateral septal nucleus, nucleus accumbens, bed nucleus of the stria terminalis, as well as the medial preoptic area and the parvocellular paraventricular nucleus of the hypothalamus. The results demonstrate that the posterolateral hypothalamus of the rat contains two regions with specific cardiovascular function and highly organized connections with diencephalic, forebrain, and cortical structures.

Functional and anatomical organization of cardiovascular pressor and depressor sites in the lateral hypothalamic area: I. Descending projections.

The present study describes the anatomical organization of projections from functionally defined cell groups of the lateral hypothalamic area. Cardiovascular pressor and depressor sites were identified following microinjection (5-50 nl) of 0.01-1.0 M L-glutamate or D,L-homocysteate into the anesthetized rat. Subsequent injections of Phaseolus vulgaris-leucoagglutinin (PHA-L) or wheat germ agglutinin-horseradish peroxidase (WGA-HRP) were made into pressor or depressor sites and their connections with the brainstem and spinal cord were traced. Decreases in blood pressure (10-45 mmHg) and heart rate (20-70 bpm) were elicited from tuberal (LHAt) and posterior (LHAp) regions of the lateral hypothalamic area (LHA). Depressor neurons in the LHAt have descending projections to the central gray, dorsal and median raphe nuclei, pedunculopontine tegmental nucleus, pontine reticular formation, medial and lateral parabrachial nuclei, laterodorsal tegmental region, and medullary reticular formation including the region of the lateral tegmental field, nucleus ambigous, and rostrocaudal ventral lateral medulla. In contrast, descending projections from depressor neurons in the LHAp have dense terminal fields in the rostral, middle, and commissural portions of the nucleus of the solitary tract and the lateral tegmental field as well as the ventrolateral central gray, pedunculopontine tegmental nucleus, and medial and lateral parabrachial nuclei. Both the LHAt and LHAp have light projections to the intermediate region of the cervical and thoracic spinal cord. Increases in blood pressure (10-40 mmHg) and heart rate (20-70 bpm) were elicited almost exclusively from neurons located medial to the LHAt and LHAp in a region surrounding the fornix, termed the perifornical area (PFA). Pressor cells in the PFA have descending projections to the central gray, dorsal and median raphe nuclei, laterodorsal tegmental nucleus, and Barrington's nucleus as well as a light projection to the commissural portion of the nucleus of the solitary tract and the intermediate region of the cervical and thoracic spinal cord. The retrograde labeling observed in the WGA-HRP studies indicates that cells in most terminal fields have reciprocal projections to the pressor and depressor regions of the LHA. The results demonstrate that groups of neurons in the lateral hypothalamus with specific cardiovascular function have differential projections to the brain stem.

Organization of visceral and limbic connections in the insular cortex of the rat.

The anterograde and retrograde transport of horseradish peroxidase was used to study the anatomical organization of visceral and limbic terminal fields in the insular cortex. Following injections into the ventroposterolateral parvicellular (VPLpc) and ventroposteromedial parvicellular (VPMpc) visceral relay nuclei of the thalamus, dense anterograde and retrograde labeling was present in the posterior granular and dysgranular insular cortices, respectively. The parabrachial nucleus had extensive connections with the posterior dysgranular cortex and to a lesser degree with the anterior dysgranular and granular cortices. In contrast, injections into the medial prefrontal cortex and mediodorsal nucleus of the thalamus resulted in dense anterograde and retrograde labeling primarily in the anterior agranular cortex, whereas injections in the amygdala resulted in axonal labeling in the agranular and dysgranular insular cortices. Injections into the lateral hypothalamic area resulted in dense anterograde and retrograde labeling mainly in the agranular and dysgranular cortices and moderate to light labeling in the granular cortex. Our results indicate that ascending visceral afferents, VPLpc, VPMpc, and parabrachial nuclei, are topographically organized in the granular and dysgranular fields of the insular cortex, whereas the agranular cortex appears to receive highly integrated limbic afferents from the infralimbic cortex and the mediodorsal nucleus of the thalamus. Although these visceral and limbic inputs to the insular cortex are segregated for the most part into different longitudinally oriented strips of cortex, limbic input from the lateral hypothalamic area and the amygdala, which have extensive autonomic as well as limbic connections, are more diffusely distributed over the different regions of the insular cortex. This organization may subserve a role for the insular cortex in integration of autonomic response with ongoing behaviour and emotion.

Topography and synaptology of mamillary body projections to the mesencephalon and pons in the rat.

The anterograde and retrograde transport of horseradish peroxidase conjugated to wheat germ agglutinin (WGA-HRP) was used to study the anatomical organization of descending projections from the mamillary body (MB) to the mesencephalon and pons at light and electron microscopic levels. Injections of WGA-HRP into the medial mamillary nucleus resulted in dense anterograde and retrograde labeling in the ventral tegmental nucleus, while injections in the lateral mamillary nucleus resulted in dense anterograde labeling in the dorsal tegmental nucleus pars dorsalis and dense anterograde and retrograde labeling in the pars ventralis of the dorsal tegmental nucleus. Anterogradely labeled fibers in the mamillotegmental tract diverged from the principal mamillary tract in an extensive dorsocaudally oriented swath of axons which extended to the dorsal and ventral tegmental nuclei, and numerous axons turned sharply ventrally and rostrally to terminate topographically in the dorsomedial nucleus reticularis tegmenti pontis and rostromedial pontine nuclei. The anterograde labeling in these two precerebellar relay nuclei was distributed near the midline such that projections from the lateral mamillary nucleus terminated mainly dorsomedial to the terminal fields of projections from the medial mamillary nucleus. In the dorsal and ventral tegmental nuclei, labeled axon terminals contained round synaptic vesicles and formed asymmetric synaptic junctions primarily with small diameter dendrites and to a lesser extent with neuronal somata. A few labeled terminals contained pleomorphic vesicles and formed symmetric synaptic junctions with dendrites and neuronal somata. Labeled axon terminals were also frequently found in synaptic contact with retrogradely labeled dendrites and neuronal somata in the dorsal and ventral tegmental nuclei. These findings indicate that neurons in the dorsal and ventral tegmental nuclei are reciprocally connected with MB projection neurons. In the nucleus reticularis tegmenti pontis and medial pontine nuclei, labeled axon terminals contained round synaptic vesicles and formed asymmetric synaptic junctions primarily with small diameter dendrites. The present study demonstrates that projections from the medial and lateral nuclei of the MB are topographically organized in the mesencephalon and pons. The synaptic morphology of mamillotegmental projections suggests that they may have excitatory influences primarily on the distal dendrites of neurons in these brain regions.