Lateral Paragigantocellular Reticular Nuclei Research Articles

Effects of stress contagion on anxiogenic- and orofacial inflammatory pain-like behaviors with brain activation in mice.

The conditions of stress contagion are induced in bystanders without direct experiences of stressful events. This study determined the effects of stress contagion on masseter muscle nociception in mice. Stress contagion was developed in the bystanders after cohabitating with a conspecific mouse subjected to social defeat stress for 10 days. On Day 11, stress contagion increased anxiety- and orofacial inflammatory pain-like behaviors. The c-Fos and FosB immunoreactivities evoked by masseter muscle stimulation were increased in the upper cervical spinal cord, while c-Fos expressions were increased in the rostral ventromedial medulla, including the lateral paragigantocellular reticular nucleus and nucleus raphe magnus in stress contagion mice. The level of serotonin in the rostral ventromedial medulla was increased under stress contagion, while the number of serotonin positive cells was increased in the lateral paragigantocellular reticular nucleus. Stress contagion increased c-Fos and FosB expressions in the anterior cingulate cortex and insular cortex, both of which were positively correlated with orofacial inflammatory pain-like behaviors. The level of brain-derived neurotrophic factor was increased in the insular cortex under stress contagion. These results indicate that stress contagion can cause neural changes in the brain, resulting in increased masseter muscle nociception, as seen in social defeat stress mice.

Differential innervation of superficial versus deep laminae of the dorsal horn by bulbo-spinal serotonergic pathways in the rat

Convergent data showed that bulbo-spinal serotonergic projections exert complex modulatory influences on nociceptive signaling within the dorsal horn. These neurons are located in the B3 area which comprises the median raphe magnus (RMg) and the lateral paragigantocellular reticular (LPGi) nuclei. Because LPGi 5-HT neurons differ from RMg 5-HT neurons regarding both their respective electrophysiological properties and responses to noxious stimuli, we used anatomical approaches for further characterization of the respective spinal projections of LPGi versus RMg 5-HT neuron subgroups.Adult Sprague-Dawley rats were stereotaxically injected into the RMg or the LPGi with the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L). The precise location of injection sites and RMg vs LPGi spinal projections into the different dorsal horn laminae were visualized by PHA-L immunolabeling. Double immunofluorescent labeling of PHA-L and the serotonin transporter (5-HTT) allowed detection of serotonergic fibers among bulbo-spinal projections.Anterograde tracing showed that RMg neurons project preferentially into the deep laminae V-VI whereas LPGi neuron projections are confined to the superficial laminae I-II of the ipsilateral dorsal horn. All along the spinal cord, double-labeled PHA-L/5-HTT immunoreactive fibers, which represent only 5–15% of all PHA-L-immunoreactive projections, exhibit the same differential locations depending on their origin in the RMg versus the LPGi.The clear-cut distinction between dorsal horn laminae receiving bulbo-spinal serotonergic projections from the RMg versus the LPGi provides further anatomical support to the idea that the descending serotonergic pathways issued from these two bulbar nuclei might exert different modulatory influences on the spinal relay of pain signaling neuronal pathways.

Terminations of reticulospinal fibers originating from the gigantocellular reticular formation in the mouse spinal cord.

The present study investigated the projections of the gigantocellular reticular nucleus (Gi) and its neighbors--the dorsal paragigantocellular reticular nucleus (DPGi), the alpha/ventral part of the gigantocellular reticular nucleus (GiA/V), and the lateral paragigantocellular reticular nucleus (LPGi)--to the mouse spinal cord by injecting the anterograde tracer biotinylated dextran amine (BDA) into the Gi, DPGi, GiA/GiV, and LPGi. The Gi projected to the entire spinal cord bilaterally with an ipsilateral predominance. Its fibers traveled in both the ventral and lateral funiculi with a greater presence in the ventral funiculus. As the fibers descended in the spinal cord, their density in the lateral funiculus increased. The terminals were present mainly in laminae 7-10 with a dorsolateral expansion caudally. In the lumbar and sacral cord, a considerable number of terminals were also present in laminae 5 and 6. Contralateral fibers shared a similar pattern to their ipsilateral counterparts and some fibers were seen to cross the midline. Fibers arising from the DPGi were similarly distributed in the spinal cord except that there was no dorsolateral expansion in the lumbar and sacral segments and there were fewer fiber terminals. Fibers arising from GiA/V predominantly traveled in the ventral and lateral funiculi ipsilaterally. Ipsilaterally, the density of fibers in the ventral funiculus decreased along the rostrocaudal axis, whereas the density of fibers in the lateral funiculus increased. They terminate mainly in the medial ventral horn and lamina 10 with a smaller number of fibers in the dorsal horn. Fibers arising from the LPGi traveled in both the ventral and lateral funiculi and the density of these fibers in the ventral and lateral funiculi decreased dramatically in the lumbar and sacral segments. Their terminals were present in the ventral horn with a large portion of them terminating in the motor neuron columns. The present study is the first demonstration of the termination pattern of fibers arising from the Gi, DPGi, GiA/GiV, and LPGi in the mouse spinal cord. It provides an anatomical foundation for those who are conducting spinal cord injury and locomotion related research.

Inhibition of cardiac baroreflex by noxious thermal stimuli: A key role for lateral paragigantocellular serotonergic cells

The present study was designed to identify the neuronal mechanisms causing cardiac baroreflex inhibition associated with thermal nociception in rats. Under urethane-anesthesia, noxious thermal stimuli ⩾48 °C were found to inhibit the cardiac baroreflex, whereas noxious stimuli ⩽46 °C had no effect. Using double immunohistochemical labeling, noxious stimuli ⩾48 °C were found to evoke primarily a strong expression of Fos protein (Fos) encoded by c-fos gene in serotonergic neurons of lateral paragigantocellular reticular nucleus (LPGi). Noxious stimuli ⩽46 °C did not evoke Fos expression in any serotonergic neurons of the brainstem. Local blockade of neuronal activity by bilateral microinjections of fluorescent muscimol (a GABA A receptor agonist tagged with a fluorophore that allowed visualization of the injections) into both the LPGi and the raphe magnus nucleus prevented the inhibitory effect of noxious stimuli ⩾48 °C on the cardiac baroreflex. Bilateral microinjections of granisetron (a 5-HT 3 antagonist) within the nucleus tractus solitarius also prevented the inhibition of cardiac baroreflex elicited by noxious stimuli ⩾48 °C. These results show that activation of serotonergic cells in the LPGi is critical to trigger nucleus tractus solitarius-mediated cardiac baroreflex inhibition elicited by intense thermal noxious stimuli.

Development of the Vestibular Apparatus and Central Vestibular Connections in a Wallaby (Macropus eugenii)

We have studied the early development of the vestibular apparatus and its central connections in the tammar wallaby (Macropus eugenii) in order to determine whether the vestibular system anatomy is sufficiently mature at birth to assist in climbing to the pouch. Structural development was studied with the aid of hematoxylin and eosin stained sections and immunoreactivity for GAP-43, whereas the development of vestibular system connections was examined by carbocyanine dye tracing. At the time of birth, the otocyst has distinct utricle, saccule and semicircular canals with immature sensory regions receiving innervation by GAP-43 immunoreactive fibers. Vestibular nerve fibers can be traced into the brainstem to the developing vestibular nuclei, which are not yet cytoarchitectonically distinct. The vestibular nuclei do not contribute direct projections to the lower cervical spinal cord at birth; most bulbospinal projections in the newborn appear to be derived bilaterally from the gigantocellular, lateral paragigantocellular reticular and ventral medullary nuclei. A substantial bilateral projection to the vestibular ganglion and apparatus from the region of the gigantocellular and lateral paragigantocellular nuclei was seen at birth, but not in subsequent ages. This is similar to a projection seen in newborn Ameridelphians. By postnatal day (P) 5, the vestibular apparatus had extensive projections to all vestibular nuclei and neurons projecting in the lateral vestibulospinal tract could be identified in the lateral vestibular nucleus. Cytoarchitectonic differentiation of the vestibular nuclei proceeded over the next 3 to 4 weeks with the emergence of discrete parvicellular and magnocellular components of the medial vestibular nucleus by P19. GAP-43 immunoreactivity stayed high in the lateral vestibulospinal tract for several months after birth, suggesting that the development of this tract followed a prolonged timecourse. Our findings indicate that central and peripheral connections of the vestibular ganglion are present at birth, but that there is no direct projection from the vestibular nuclei to the cervical spinal cord until P5. Nevertheless, the possibility remains that an indirect projection between the vestibular nuclei and the medial reticular formation is present at birth and mediates control of the climb.

Critical role of B3 serotonergic cells in baroreflex inhibition during the defense reaction triggered by dorsal periaqueductal gray stimulation

The present study was designed to identify the serotonergic pathway causing baroreflex inhibition associated with the defense reaction in rats. Under conditions that produce physiological responses typical of the defense reaction, electrical stimulation of the dorsal periaqueductal gray (dPAG) was found to double c-Fos immunoreactive serotonergic neurons within the mid-rostrocaudal extent of the B3 group (which comprises the raphe magnus and the lateral paragigantocellular reticular nuclei) in anesthetized rats. Local blockade of neuronal activity by microinjection of muscimol (a GABA(A) receptor agonist) directly into the B3 region prevented the inhibitory effect of dPAG activation on the cardiac baroreflex. Conversely, neuron activation by local application of D,L-homocysteic acid into B3 region caused baroreflex inhibition that was suppressed by microinjection of granisetron (a 5-HT(3) antagonist) into the nucleus tractus solitarius. These results show that activation of serotonergic cells in the mid-portion of B3 group is critical to trigger baroreflex inhibition occurring during the defense reaction evoked by dPAG stimulation.

Localization of the neurons active during paradoxical (REM) sleep and projecting to the locus coeruleus noradrenergic neurons in the rat

Locus coeruleus (LC) noradrenergic neurons are active during wakefulness, slow their discharge rate during slow wave sleep, and stop firing during paradoxical sleep (PS). A large body of data indicates that their inactivation during PS is due to a tonic GABAergic inhibition. To localize the neurons responsible for such inhibition, we first examined the distribution of retrogradely and Fos double-immunostained neurons following cholera toxin b subunit (CTb) injection in the LC of control rats, rats selectively deprived of PS for 3 days, and rats allowed to recover for 3 hours from such deprivation. We found a significant number of CTb/Fos double-labeled cells only in the recovery group. The largest number of CTb/Fos double-labeled cells was found in the dorsal paragigantocellular reticular nucleus (DPGi). It indeed contained 19% of the CTb/Fos double-labeled neurons, whereas the ventrolateral periaqueductal gray (vlPAG) contained 18.3% of these neurons, the lateral paragigantocellular reticular nucleus (LPGi) 15%, the lateral hypothalamic area 9%, the lateral PAG 6.7%, and the rostral PAG 6%. In addition, CTb/Fos double-labeled cells constituted 43% of all the singly CTb-labeled cells counted in the DPGi compared with 29% for the LPGi, 18% for the rostral PAG, and 10% or less for the other structures. Although all these populations of CTb/Fos double-labeled neurons could be GABAergic and tonically inhibit LC neurons during PS, our results indicate that neurons from the DPGi constitute the best candidate for this role.

Cholinergic and noncholinergic brainstem neurons expressing Fos after paradoxical (REM) sleep deprivation and recovery

It is well accepted that populations of neurons responsible for the onset and maintenance of paradoxical sleep (PS) are restricted to the brainstem. To localize the structures involved and to reexamine the role of mesopontine cholinergic neurons, we compared the distribution of Fos- and choline acetyltransferase-labelled neurons in the brainstem of control rats, rats selectively deprived of PS for approximately 72 h and rats allowed to recover from such deprivation. Only a few cholinergic neurons from the laterodorsal (LDTg) and pedunculopontine tegmental nuclei were Fos-labelled after PS recovery. In contrast, a large number of noncholinergic Fos-labelled cells positively correlated with the percentage of time spent in PS was observed in the LDTg, sublaterodorsal, alpha and ventral gigantocellular reticular nuclei, structures known to contain neurons specifically active during PS. In addition, a large number of Fos-labelled cells were seen after PS rebound in the lateral, ventrolateral and dorsal periaqueductal grey, dorsal and lateral paragigantocellular reticular nuclei and the nucleus raphe obscurus. Interestingly, half of the cells in the latter nucleus were immunoreactive to choline acetyltransferase. In contrast to the well-accepted hypothesis, our results strongly suggest that neurons active during PS, recorded in the mesopontine cholinergic nuclei, are in the great majority noncholinergic. Our findings further demonstrate that many brainstem structures not previously identified as containing neurons active during PS contain cholinergic or noncholinergic neurons active during PS, and these structures may therefore play a key role during this state. Altogether, our results open a new avenue of research to identify the specific role of the populations of neurons revealed, their interrelations and their neurochemical identity.

The rostral ventromedial medulla mediates the facilitatory effect of microinjected orphanin FQ in the periaqueductal gray on spinal nociceptive transmission in rats

Single unit extracellular recordings from spinal dorsal horn neurons were obtained with glass micropipettes in pentobarbital-anesthetized rats. A total of 50 wide dynamic range (WDR) neurons were studied in 25 rats. Microinjection of orphanin FQ (OFQ, 0.1 μg/0.1 μl) (a potent endogenous ligand of the opioid receptor-like receptor (ORL-1)) into the ipsilateral ventrolateral parts of periaqueductal gray (vlPAG) significantly increased C-response and post-discharge activity in most of the WDR neurons. Pre-microinjection of lidocaine (4%) into the nucleus raphe magnus (NRM) (0.5 μl), ipsilateral nucleus reticularis gigantocellularis (NGC) (0.6 μl), or nucleus gigantocellularis pars alpha (NGCα) and nucleus reticularis paragigantocellularis lateralis (NPGL) (0.5 μl) markedly reduced intra-vlPAG microinjection of OFQ-induced facilitatory effects on nociceptive responses of WDR neurons. Furthermore, if the NRM and ipsilateral NGC were simultaneously pre-microinjected with lidocaine, the intra-vlPAG microinjection of OFQ-induced facilitation on nociceptive responses of WDR neurons was eliminated. Also, a similar effect was observed when all the NRM, ipsilateral NGC, NGCα and NPGL were blocked with lidocaine. No significant effect on nociceptive responses of WDR neurons per se was found after blocking the NRM, ipsilateral NGC, NGCα/NPGL, or all the NRM, ipsilateral NGC, and NGCα/NPGL with lidocaine. These results indicate that (1) the facilitatory effect evoked by microinjection of OFQ into the vlPAG on nociceptive responses of WDR neurons in the spinal dorsal horn is primarily mediated by the NRM and ipsilateral NGC; (2) the NRM, ipsilateral NGC, and NGCα/NPGL do not mediate tonic descending inhibition of the spinal dorsal horn neurons.

Pain pathways and parabrachial circuits in the rat.

This review presents a schematic attempt to classify the major pain pathways, based on the results of recent studies in our laboratory, with a special emphasis on the parabrachial system. Our view is based on results from experiments in the rat, using very small iontophoretic injections of anterograde tracers. As illustrated in this report, we have found a very dramatic difference between ascending projections originating from deep laminae compared with those arising from lamina I of the dorsal horn. We propose three main pain systems and discuss their functional-anatomical relationships. The first system is centred on the projection from deep laminae to three caudal reticular areas - the lateral reticular nucleus (LRN), the subnucleus reticularis dorsalis (SRD) and the gigantocellular lateral paragigantocellular reticular nuclei (NGc) - and the parabrachial internal lateral subnucleus (PBil). The second system is centred on the projection from lamina I to the ventral posterolateral nucleus (VPL), the ventral posteromedial (VPM), the posterior nuclear group (Po) and triangular posterior nucleus (PoT) of the thalamus. The third system is centred on the projection from lamina I to the lateral parabrachial area. We also present the four main projections from the latter area to the extended amygdala, the hypothalamus, the periaqueductal grey matter (PAG), and the ventrolateral medulla (VLM), and their involvement in emotional and autonomic (homeostatic) aspects of pain.

Involvement of nitric oxide in regulation of the medullary respiratory rhythm in neonatal rats

The NADPH-diaphorase (as a neuronal NO-synthase) reactivity in the medullary structures of the respiratory rhythm (RR) generator and the role of NO in the regulation of respiratory activity in the phrenic nerve of artificially superfused semi-isolated medulla-spinal cord preparations were investigated in newborn rats. NADPH-diaphorase positive neurons were found in all nuclei of both dorsal and ventral respiratory groups of neurons. The maximal density of stained cells was present within the rostral part of the ventrolateral medulla (VLM), in the region of the lateral paragigantocellular reticular nucleus. It was found that endogenous NO mediates the mechanism of tonic inhibitory control of the RR frequency located in the rostral VLM under normal and hypoxic conditions, and appears to be involved in generation of the basic RR by the more caudal structures of VLM. It was shown that NO biosynthesis mediates the effect of NMDA receptors activation on the RR.

Brainstem microstimulation activates sympathetic fibers in pudendal nerve motor branch.

A bilateral spino-bulbo-spinal circuit conveys information from/to the male urogenital tract and perineal muscles. This is the first electrophysiological report of another descending pathway, one which conveys output from the medullary reticular formation (MRF) to activate postganglionic sympathetic fibers contained within the motor branch of the pudendal nerve (PudM). In anesthetized rats, long latency (> 150 ms) discharges were elicited in the PudM following ipsilateral or contralateral microstimulation of the MRF. These firing bursts were not observed in rats after sectioning the lower lumbar sympathetic trunk. The most robust activation was observed when neurons in or near the lateral paragigantocellular reticular nucleus were microstimulated bilaterally. Urogenital dysfunction that occurs following severe spinal cord injury probably results from disrupting these and other supraspinal circuits.

Sulfated cholecystokinin octapeptide in the rat: pontomedullary distribution and modulation of the respiratory pattern

We performed anatomical and physiological studies to determine the site and actions of sulfated cholecystokinin octapeptide (CCK8-S) on breathing. Peptide locations were determined by combined immunodetection of CCK8-S- containing synaptic varicosities and retrograde labeling of medullary neurons projecting to the ventral respiratory group. Retrogradely labeled neurons and CCK8-S immunolabeled varicosities overlapped within the nuclei of the solitary tract, ventral respiratory group, and the Kolliker-Fuse nucleus. Additional CCK8-S immunoreactive terminals were located in the rostroventrolateral medullary reticular nucleus, lateral paragigantocellular reticular nucleus, and the caudal pontine reticular nucleus. The respiratory effects of CCK8-S, which binds to CCK(A) and CCK(B) receptors, were examined by intravenous injection in adult rats and by bath application in the in vitro neonatal rat brainstem - spinal cord preparation. CCK8-S produced an increase in the mean amplitude of diaphragmatic electromyogram (EMG) of 28 +/- 35% (SD) and a decrease in mean respiratory interval of 13 +/- 4% in vivo. In vitro, CCK8-S significantly increased inspiratory duration and decreased respiratory interval, primarily by shortening expiratory duration. CCK8-unsulfated, a specific agonist for CCK(B) receptors, did not produce these effects. CCK8-S effects in the in vitro preparation were partially blocked by the CCK receptor antagonist lorglumide (final bath concentration 600 nM). These results suggest that CCK8-S modulates the respiratory rhythm via CCK(A) receptors within one or more medullary or pontine respiratory groups in both neonatal and adult rats.

CNS innervation of vagal preganglionic neurons controlling peripheral airways: a transneuronal labeling study using pseudorabies virus

The CNS cell groups that project to vagal preganglionic neurons which innervate the most distal part of the airways were identified by the viral retrograde transneuronal labeling method. Pseudorabies virus (PRV) was injected into the lung parenchyma of C8 spinal rats and after 5 days survival, brain tissue sections from these animals were processed for immunohistochemical detection of PRV. Retrogradely labeled parasympathetic preganglionic cells (first-order neurons) were seen mainly in the ventral medulla oblongata: the compact portion of the nucleus ambiguus and the area ventral to it. Occasionally, a few labeled cells were seen within the rostral part of the dorsal vagal nucleus. This labeling pattern correlated well with the retrograde cell body labeling seen following cholera toxin β-subunit (CT-b) injections in the lung parenchyma. PRV transneuronally labeled neurons (second-order and/or presumed third-order neurons) were found throughout the CNS with the characteristic labeling in the brainstem. Labeled neurons were identified along and just beneath the ventral medullary surface, and in nearby areas: the parapyramidal, retrotrapezoid, gigantocellular and lateral paragigantocellular reticular nuclei, as well as the caudal raphe nuclei (raphe pallidus, obscurus, and magnus). Several nucleus tractus solitarius (nTS) regions contained labeled cells including the commissural, medial, and ventrolateral nTS subnuclei. The A5 cell group and a small number of locus coeruleus neurons were also labeled. PRV-infected neurons were present in the Kölliker–Fuse and Barrington's nuclei. In the mesencephalon, neurons within the ventral periventricular gray matter were labeled. Labeling was present in the dorsal, lateral and paraventricular hypothalamic nuclei, and within the amygdaloid complex. In summary, the parasympathetic preganglionic neurons that innervate the peripheral airways are controlled by networks of lower brainstem and suprapontine neurons that lie in the same regions known to be involved in central regulation of autonomic functions.

Sulfated cholecystokinin octapeptide in the rat: pontomedullary distribution and modulation of the respiratory pattern

We performed anatomical and physiological studies to determine the site and actions of sulfated cholecystokinin octapeptide (CCK8-S) on breathing. Peptide locations were determined by combined immunodetection of CCK8-S- containing synaptic varicosities and retrograde labeling of medullary neurons projecting to the ventral respiratory group. Retrogradely labeled neurons and CCK8-S immunolabeled varicosities overlapped within the nuclei of the solitary tract, ventral respiratory group, and the Kolliker-Fuse nucleus. Additional CCK8-S immunoreactive terminals were located in the rostroventrolateral medullary reticular nucleus, lateral paragigantocellular reticular nucleus, and the caudal pontine reticular nucleus. The respiratory effects of CCK8-S, which binds to CCK(A) and CCK(B) receptors, were examined by intravenous injection in adult rats and by bath application in the in vitro neonatal rat brainstem - spinal cord preparation. CCK8-S produced an increase in the mean amplitude of diaphragmatic electromyogram (EMG) of 28 +/- 35% (SD) and a decrease in mean respiratory interval of 13 +/- 4% in vivo. In vitro, CCK8-S significantly increased inspiratory duration and decreased respiratory interval, primarily by shortening expiratory duration. CCK8-unsulfated, a specific agonist for CCK(B) receptors, did not produce these effects. CCK8-S effects in the in vitro preparation were partially blocked by the CCK receptor antagonist lorglumide (final bath concentration 600 nM). These results suggest that CCK8-S modulates the respiratory rhythm via CCK(A) receptors within one or more medullary or pontine respiratory groups in both neonatal and adult rats.

Brainstem microstimulation differentially inhibits pudendal motoneuron reflex inputs.

Sensory information from the genitourinary tract is transmitted to perineal muscle reflex circuits in the spinal cord and to the medullary reticular formation (MRF). This is the first report of a functional descending connection between neurons in the MRF and sensory inputs of the pudendal motoneuron reflex circuitry. In anesthetized rats, microstimulation of the MRF produced a decrease in amplitude and increase in latency of pudendal motoneuron reflex discharges (PMRD) elicited by stimulation of the dorsal nerve of the penis. No effects on pelvic nerve-elicited PMRD were found. The most robust reflex depression was observed when the lateral paragigantocellular reticular nucleus was microstimulated bilaterally. Reproductive abnormalities that occur following severe spinal cord injury probably result from disruption of these descending connections.

Afferent and efferent connections of the ventrolateral tegmental area in the rat.

The present study examined the organization of afferent and efferent connections of the rat ventrolateral tegmental area (VLTg) by employing the retrograde and anterograde axonal transport of Fluorogold and Phaseolus vulgaris-leucoagglutinin, respectively. Our interest was focused on whether the anatomical connections of the VLTg would provide evidence as to the involvement of this reticular area in audiomotor behavior. Our retrograde experiments revealed that minor inputs to the VLTg arise in various telencephalic structures, including the cerebral cortex. Stronger projections originate in the lateral preoptic area, the zona incerta, the nucleus of the posterior commissure and some other thalamic areas, the lateral substantia nigra, the deep layers of the superior colliculus, the dorsal and lateral central gray, the deep mesencephalic nucleus, the paralemniscal zone, the intercollicular nucleus, the external cortex of the inferior colliculus, the oral and caudal pontine reticular nucleus, the deep cerebellar nuclei, the gigantocellular and lateral paragigantocellular reticular nuclei, the prepositus hypoglossal nucleus, the spinal trigeminal nuclei, and the intermediate layers of the spinal cord. Most importantly, we disclosed strong auditory afferents arising in the dorsal and ventral cochlear nuclei and in the cochlear root nucleus. The efferent projections of the VLTg were found to be less widespread. Telencephalic structures do not receive any input from the VLTg. Moderate projections were seen to diencephalic reticular areas, the zona incerta, the nucleus of the posterior commissure, and to various other thalamic areas. The major VLTg projections terminate in the deep layers of the superior colliculus, the deep mesencephalic nucleus, the intercollicular nucleus and external cortex of the inferior colliculus, the oral and caudal pontine reticular nucleus, the gigantocellular and lateral paragigantocellular reticular nuclei, and in the medial column of the facial nucleus. From our data, we conclude that the VLTg might play a role in sensorimotor behavior.

CNS sites involved in sympathetic and parasympathetic control of the pancreas: a viral tracing study

The viral transneuronal tracing method was used to identify the CNS cell groups that regulate the parasympathetic and sympathetic outflow systems of the pancreas. Pseudorabies virus (PRV) was injected into the pancreas of vagotomized rats and after 6 days survival, the pattern of transneuronal labeling in the CNS sympathetic regulatory regions was determined. The converse experiment was performed in order to elucidate the central parasympathetic cell groups that regulate the pancreas. Immunohistochemical methods were used to identify putative neuropeptide- and catecholamine-containing CNS neurons involved in these regulatory circuits. The major finding of this study indicates that five brain regions, viz., paraventricular hypothalamic nucleus, perifornical hypothalamic region, A5 catecholamine cell group, rostral ventrolateral medulla, and lateral paragigantocellular reticular nucleus, contain a considerable amount of overlap in cell body labeling. In addition, the ventrolateral part of the periaqueductal gray matter and gigantocellular reticular nucleus, ventral part also showed a similar overlap, but the numbers of neurons found in these areas were considerably lower than the five major regions. These data suggest that these brain regions may provide parallel and possibly redundant, autonomic pathways affecting glucagon and adrenaline release.

Afferent projections to the rat nuclei raphe magnus, raphe pallidus and reticularis gigantocellularis pars α demonstrated by iontophoretic application of choleratoxin (subunit b)

The aim of the present study was to identify the specific afferent projections to the rostral and caudal nucleus raphe magnus, the gigantocellular reticular nucleus pars α and the rostral nucleus raphe pallidus. For this purpose, small iontophoretic injections of the sensitive retrograde tracer choleratoxin (subunit b) were made in each of these structures. In agreement with previous retrograde studies, after all injection sites, a substantial to large number of labeled neurons were observed in the dorsal hypothalamic area and dorsolateral and ventrolateral parts of the periaqueductal gray, and a small to moderate number were found in the lateral preoptic area, bed nucleus of the stria terminalis, paraventricular hypothalamic nucleus, central nucleus of the amygdala, lateral hypothalamic area, parafascicular area, parabrachial nuclei, subcoeruleus area and parvocellular reticular nucleus. In addition, depending on the nucleus injected, we observed a variable number of retrogradely labeled cells in other regions. After injections in the rostral nucleus raphe magnus, a large number of labeled cells were seen in the prelimbic, infralimbic, medial and lateral precentral cortices and the dorsal part of the periaqueductal gray. In contrast, after injections in the other nuclei, fewer cells were localized in these structures. Following raphe pallidus injections, a substantial to large number of labeled cells were observed in the medial preoptic area, median preoptic nucleus, ventromedial part of the periaqueductal gray, Kölliker-Fuse and lateral paragigantocellular reticular nuclei. Following injections in the other areas, a small to moderate number of cells appeared. After gigantocellular reticular pars α injections, a very large and substantial number of labeled neurons were found in the deep mesencephalic reticular formation and oral pontine reticular nucleus, respectively. After the other injections, fewer cells were seen. Following rostral raphe magnus or raphe pallidus injections, a substantial number of labeled cells were observed in the insular and perirhinal cortices. Following caudal raphe magnus or gigantocellular reticular pars α injections, fewer cells were found. After raphe magnus or gigantocellular reticular pars α injections, a moderate to substantial number of cells were localized in the fields of Forel, lateral habenular nucleus and ventral caudal pontine reticular nucleus. Following raphe pallidus injections, only a small number of cells were seen. Our data indicate that the rostral and caudal parts of the nucleus raphe magnus, the gigantocellular reticular nucleus pars α and the nucleus raphe pallidus receive afferents of comparable strength from a large number of structures. In addition, a number of other afferents give rise to stronger inputs to one or two of the four nuclei studied. Such differential inputs might be directed to populations of neurons with different physiological roles previously recorded specifically in these nuclei.

Physiological identification of pontomedullary serotonergic neurons in the rat.

Spinal serotonin is derived entirely from bulbar sources and plays an important role in spinal modulatory processes, including pain modulation. Establishing the electrophysiological properties of SEROTONERGIC bulbospinal neurons in the pontomedullary raphe and reticular formation is critical to understanding the physiological role of serotonin in the spinal cord. Neurons were characterized by their responses to noxious stimulation and their background discharge pattern in the lightly anesthetized rat. Characterized cells were intracellularly labeled with Neurobiotin, which was visualized with a Texas Red fluorophore. Sections containing the labeled cells were processed for serotonin immunocytochemistry with the use of a Bodipy fluorophore. Forty-seven intracellularly labeled cells were tested for serotonin immunoreactivity. The labeled neurons were located in raphe magnus, the nucleus reticularis magnocellularis, and the adjacent reticular and raphe nuclei at levels from the inferior olivary complex to the superior olivary complex. SEROTONERGIC cells were located in the raphe nuclei, in nucleus reticularis magnocellularis pars alpha, and in nucleus reticularis magnocellularis pars beta or nucleus reticularis gigantocellularis. Thirteen intracellularly labeled cells contained serotonin immunoreactivity. The background discharge rate of SEROTONERGIC cells average 1.8 Hz (range: 0.5-3.1 Hz). Discharge was steady and without sustained pauses or bursts in firing. Most serotonin-immunoreactive cells were unaffected or slightly excited by pinch and were unaffected by noxious heat. Three SEROTONERGIC cells were weakly excited by both noxious pinch and heat, whereas two SEROTONERGIC cells were briefly inhibited by these stimuli. Cells that lacked serotonin immunoreactivity were heterogeneous and included ON, OFF, and NEUTRAL cells. Nonserotonergic cells differed from SEROTONERGIC cells in having an irregular discharge pattern and/or a high mean discharge rate. A linear discriminant function, employing background discharge characteristics as independent variables, was calculated that successfully classified 13 of 13 SEROTONERGIC and 32 of 33 nonserotonergic neurons. The probability of misclassification with the use of this discriminant function was estimated to be < 10%. Employing the discriminant function on a test group of cells whose immunochemical content was unknown revealed a population of SEROTONERGIC-LIKE cells that resembled the labeled SEROTONERGIC cells in background discharge pattern, response to noxious stimulation, and nuclear location. The discharge of pontomedullary SEROTONERGIC neurons is slow and steady, suggesting that these neurons may have a role in the tonic, rather than phasic, modulation of spinal processes.