Parvicellular Reticular Nucleus Research Articles

Brain-wide mapping of c-Fos expression in nitroglycerin-induced models of migraine

BackgroundMigraine is a neurological disorder characterized by complex, widespread, and sudden attacks with an unclear pathogenesis, particularly in chronic migraine (CM). Specific brain regions, including the insula, amygdala, thalamus, and cingulate, medial prefrontal, and anterior cingulate cortex, are commonly activated by pain stimuli in patients with CM and animal models. This study employs fluorescence microscopy optical sectioning tomography (fMOST) technology and AAV-PHP.eB whole-brain expression to map activation patterns of brain regions in CM mice, thus enhancing the understanding of CM pathogenesis and suggesting potential treatment targets.MethodsBy repeatedly administering nitroglycerin (NTG) to induce migraine-like pain in mice, a chronic migraine model (CMM) was established. Olcegepant (OLC) was then used as treatment and its effects on mechanical pain hypersensitivity and brain region activation were observed. All mice underwent mechanical withdrawal threshold, light-aversive, and elevated plus maze tests. Viral injections were administered to the mice one month prior to modelling, and brain samples were collected 2 h after the final NTG/vehicle control injection for whole-brain imaging using fMOST.ResultsIn the NTG-induced CMM, mechanical pain threshold decreased, photophobia, and anxiety-like behavior were observed, and OLC was found to improve these manifestations. fMOST whole-brain imaging results suggest that the isocortex-cerebral cortex plate region, including somatomotor areas (MO), somatosensory areas (SS), and main olfactory bulb (MOB), appears to be the most sensitive area of activation in CM (P < 0.05). Other brain regions such as the inferior colliculus (IC) and intermediate reticular nucleus (IRN) were also exhibited significant activation (P < 0.05). The improvement in migraine-like symptoms observed with OLC treatment may be related to its effects on these brain regions, particularly SS, MO, ansiform lobule (AN), IC, spinal nucleus of the trigeminal, caudal part (Sp5c), IRN, and parvicellular reticular nucleus (PARN) (P < 0.05).ConclusionsfMOST whole-brain imaging reveals c-Fos + cells in numerous brain regions. OLC improves migraine-like symptoms by modulating brain activity in some brain regions. This study demonstrates the activation of the specific brain areas in NTG-induced CMM and suggests some regions as a potential treatment mechanism according to OLC.Graphical

Dystonia-like behaviors and impaired sensory-motor integration following neurotoxic lesion of the pedunculopontine tegmental nucleus in mice.

The pedunculopontine nucleus (PPTg) is a vital interface between the basal ganglia and cerebellum, participating in modulation of the locomotion and muscle tone. Pathological changes of the PPTg have been reported in patients and animal models of dystonia, while its effect and mechanism on the phenotyping of dystonia is still unknown. In this study, a series of behavioral tests focusing on the specific deficits of dystonia were conducted for mice with bilateral and unilateral PPTg excitotoxic lesion, including the dystonia-like movements evaluation, different types of sensory-motor integrations, explorative behaviors and gait. In addition, neural dysfunctions including apoptosis, neuroinflammation, neurodegeneration and neural activation of PPTg-related motor areas in the basal ganglia, reticular formations and cerebellum were also explored. Both bilateral and unilateral lesion of the PPTg elicited dystonia-like behaviors featured by the hyperactivity of the hindlimb flexors. Moreover, proprioceptive and auditory sensory-motor integrations were impaired in bilaterally lesioned mice, while no overt alterations were found for the tactile sensory-motor integration, explorative behaviors and gait. Similar but milder behavioral deficits were found in the unilaterally lesioned mice, with an effective compensation was observed for the auditory sensory-motor integration. Histologically, no neural loss, apoptosis, neuroinflammation and neurodegeneration were found in the substantia nigra pars compacta and caudate putamen (CPu) following PPTg lesion, while reduced neural activity was found in the dorsolateral part of the CPu and striatal indirect pathway-related structures including subthalamic nucleus, globus pallidus internus and substantia nigra pars reticular. Moreover, the neural activity was decreased for the reticular formations such as pontine reticular nucleus, parvicellular reticular nucleus and gigantocellular reticular nucleus, while deep cerebellar nuclei were spared. In conclusion, lesion of the PPTg could elicit dystonia-like behaviors through its effect on the balance of the striatal pathways and the reticular formations.

Cortical Neurostimulation and N-Methyl-D-Aspartate Glutamatergic Receptor Activation in the Dysgranular Layer of the Posterior Insular Cortex Modulate Chronic Neuropathic Pain

Background and AimsThe dysgranula parts of the posterior insular cortex (PIC) stimulation (PICS) has been investigated as a new putative cortical target for nonpharmacologic therapies in patients with chronic and neuropathic pain (NP). This work investigates the neural bases of insula neurostimulation–induced antinociception and glutamatergic neurochemical mechanisms recruited by the PICS in animals with neuropathy. Materials and MethodsMale Wistar rats were submitted to the von Frey and acetone tests to assess mechanical and cold allodynia after 21 days of chronic constriction injury (CCI) of the sciatic nerve or Sham procedure (“false operated”). Either the Cascade Blue 3000 MW lysine-fixable dextran (CBD) or the biotinylated dextran amine 3000 MW (BDA) neural tract tracer was microinjected into the PIC. The electrical PICS was performed at a low frequency (20 μA, 100 Hz) for 15 seconds by a deep brain stimulation device. PIC N-methyl-D-aspartate (NMDA) receptors (NMDAR) blockade with the selective antagonist LY235959 (at 2, 4, and 8 nmol/200 nL) followed by PICS was investigated in rats with CCI. ResultsPIC sends projections to the caudal pontine reticular nucleus, alpha part of the parvicellular reticular nucleus, dorsomedial tegmental area, and secondary somatosensory cortex (S2). PICS decreased both mechanical and cold allodynia in rats with chronic NP. Blockade of NMDAR in the PIC with LY235959 at 8 nmol attenuated PICS-produced antinociception. ConclusionNeuroanatomic projections from the PIC to pontine reticular nuclei and S2 may contribute to chronic NP signaling. PICS attenuates the chronic NP, and the NMDA glutamatergic system in the PIC may be involved in PICS-induced antinociception in rodents with NP conditions.

Functional connectome of brainstem nuclei involved in autonomic, limbic, pain and sensory processing in living humans from 7 Tesla resting state fMRI

Despite remarkable advances in mapping the functional connectivity of the cortex, the functional connectivity of subcortical regions is understudied in living humans. This is the case for brainstem nuclei that control vital processes, such as autonomic, limbic, nociceptive and sensory functions. This is because of the lack of precise brainstem nuclei localization, of adequate sensitivity and resolution in the deepest brain regions, as well as of optimized processing for the brainstem. To close the gap between the cortex and the brainstem, on 20 healthy subjects, we computed a correlation-based functional connectome of 15 brainstem nuclei involved in autonomic, limbic, nociceptive, and sensory function (superior and inferior colliculi, ventral tegmental area-parabrachial pigmented nucleus complex, microcellular tegmental nucleus-prabigeminal nucleus complex, lateral and medial parabrachial nuclei, vestibular and superior olivary complex, superior and inferior medullary reticular formation, viscerosensory motor nucleus, raphe magnus, pallidus, and obscurus, and parvicellular reticular nucleus – alpha part) with the rest of the brain. Specifically, we exploited 1.1mm isotropic resolution 7 Tesla resting-state fMRI, ad-hoc coregistration and physiological noise correction strategies, and a recently developed probabilistic template of brainstem nuclei. Further, we used 2.5mm isotropic resolution resting-state fMRI data acquired on a 3 Tesla scanner to assess the translatability of our results to conventional datasets. We report highly consistent correlation coefficients across subjects, confirming available literature on autonomic, limbic, nociceptive and sensory pathways, as well as high interconnectivity within the central autonomic network and the vestibular network. Interestingly, our results showed evidence of vestibulo-autonomic interactions in line with previous work. Comparison of 7 Tesla and 3 Tesla findings showed high translatability of results to conventional settings for brainstem-cortical connectivity and good yet weaker translatability for brainstem-brainstem connectivity. The brainstem functional connectome might bring new insight in the understanding of autonomic, limbic, nociceptive and sensory function in health and disease.

The Mammalian Locus Coeruleus Complex-Consistencies and Variances in Nuclear Organization.

Descriptions of the nuclear parcellation of the locus coeruleus complex have been provided in approximately 80 mammal species spanning the phylogenetic breadth of this class. Within the mammalian rostral hindbrain, noradrenergic neurons (revealed with tyrosine hydroxylase and dopamine-ß-hydroxylase immunohistochemistry) have been observed within the periventricular grey matter (A4 and A6 nuclei) and parvicellular reticular nucleus (A5 and A7 nuclei), with the one exception to date being the tree pangolin, where no A4/A6 neurons are observed. The alphanumeric nomenclature system, developed in laboratory rodent brains, has been adapted to cover the variation observed across species. Cross-species homology is observed regarding the nuclear organization of noradrenergic neurons located in the parvicellular reticular nucleus (A5 and A7). In contrast, significant variations are observed in the organization of the A6 neurons of the locus coeruleus proper. In most mammals, the A6 is comprised of a moderate density of neurons, but in Murid rodents, primates, and megachiropteran bats, the A6 exhibits a very high density of neurons. In primates and megachiropterans, there is an additional moderate density of A6 neurons located rostromedial to the high-density portion. These variations are of importance in understanding the translation of findings in laboratory rodents to humans.

Whole-brain mapping of afferent projections to the suprachiasmatic nucleus of the tree shrew

The suprachiasmatic nucleus (SCN) is essential for the neural control of mammalian circadian timing system. The circadian activity of the SCN is modulated by its afferent projections. In the present study, we examine neuroanatomical characteristics and afferent projections of the SCN in the tree shrew (Tupaia belangeri chinensis) using immunocytochemistry and retrograde tracer Fluoro-Gold (FG). Distribution of the vasoactive intestinal peptide was present in the SCN from rostral to caudal, especially concentrated in its ventral part. FG-labeled neurons were observed in the lateral septal nucleus, septofimbrial nucleus, paraventricular thalamic nucleus, posterior hypothalamic nucleus, posterior complex of the thalamus, ventral subiculum, rostral linear nucleus of the raphe, periaqueductal gray, mesencephalic reticular formation, dorsal raphe nucleus, pedunculopontine tegmental nucleus, medial parabrachial nucleus, locus coeruleus, parvicellular reticular nucleus, intermediate reticular nucleus, and ventrolateral reticular nucleus. In summary, the morphology of the SCN in tree shrews is described from rostral to caudal. In addition, our data demonstrate for the first time that the SCN in tree shrews receives inputs from numerous brain regions in the telencephalon, diencephalon, mesencephalon, metencephalon, and myelencephalon. This comprehensive knowledge of the afferent projections of the SCN in tree shrews provides further insights into the neural organization and physiological processes of circadian rhythms.

Whole-brain monosynaptic inputs and outputs of glutamatergic neurons of the vestibular nuclei complex in mice

Vestibular nuclei complex (VN) glutamatergic neurons play a critical role in the multisensory and multimodal processing. The dysfunction of VN leads to a series of vestibular concurrent symptoms, such as disequilibrium, spatial disorientation, autonomic disorders and even emotion disorders. However, the reciprocal neural connectivity in the whole brain of VN glutamatergic neurons was incompletely understood. Here, we employed a cell-type-specific, cre-dependent, modified virus vector to retrogradely and anterogradely trace VN glutamatergic neurons in the VGLUT2-IRES-Cre mouse line. We identified and analyzed statistically the afferents and efferents of VN glutamatergic neurons in the whole brain, and also reconstructed monosynaptic inputs distribution of VN glutamatergic neurons at the three-dimensional level with the combination of a fluorescence micro-optical sectioning tomography system (fMOST). We found that VN glutamatergic neurons primarily received afferents from 57 nuclei and send efferents to 59 nuclei in the whole brain, intensively located in the brainstem and cerebellum. Projections from nuclei in the cerebellum targeting VN glutamatergic neurons mainly performed the balance control – the principal function of the vestibular system. In addition, VN glutamatergic neurons sent projections to oculomotor nucleus, trochlear nucleus and abducens nucleus dominating the eye movement. Except for the maintenance of balance, VN glutamatergic neurons were also directly connected with other functional regions, such as sleep-wake state (locus coeruleus, dorsal raphe nucleus, and laterodorsal tegmental nucleus, gigantocellular reticular nucleus, lateral paragigantocellular nucleus, periaqueductal gray, subcoeruleus nucleus, parvicellular reticular nucleus, paramedian raphe nucleus), and emotional regulation (locus coeruleus and dorsal raphe nucleus). Hence, this study revealed a comprehensive whole-brain neural connectivity of VN glutamatergic neurons and provided with a neuroanatomic foundation to further study on central vestibular circuits.

Neuroanatomical correlates of the inhibition of tremulous jaw movements in rats by a combination of memantine and Δ9 -tetrahydrocannabinol.

Memantine and marijuana smoking have been found to inhibit tremor in parkinsonian patients, although the observed effects were relatively weak. The tremorolytic effects of combinations of memantine and cannabinoids have not been studied. Here, we have evaluated the anti-tremor activity of memantine, Δ9 -tetrahydrocannabinol (THC) given alone and of their combination. The involvement of some neuroanatomical structures in the effects of the combination was evaluated. Haloperidol-induced tremulous jaw movements (TJMs) in rats were used as a model of parkinsonian-like tremor. To evaluate the role of central receptor systems in the drug effects, receptor ligands were administered locally into certain brain areas. Memantine and THC alone were without effect, although co-administration of these drugs decreased the number of haloperidol-induced jaw movements. The anti-tremor activity of the combination was antagonized (a) by injections of l-glutamate into the dorsal striatum, entopeduncular nucleus, substantia nigra pars reticulata, globus pallidus, and supratrigeminal and trigeminal motor nuclei but not into the subthalamic and cuneiform nuclei; (b) by injections of CGS 21680 into the ventrolateral striatum; and (c) by injections of bicuculline into the rostral part of the parvicellular reticular nucleus. Memantine and THC supra-additively inhibit haloperidol-induced TJMs, suggesting that co-administration of these drugs might be a new approach to the treatment of tremor. Our results identified brain areas influencing parkinsonian-like tremor in rats and can help advance the development of novel treatments for repetitive involuntary movements.

Medullary Reticular Neurons Mediate Neuropeptide Y-Induced Metabolic Inhibition and Mastication

Hypothalamic neuropeptide Y (NPY) elicits hunger responses to increase the chances of surviving starvation: an inhibition of metabolism and an increase in feeding. Here we elucidate a key central circuit mechanism through which hypothalamic NPY signals drive these hunger responses. GABAergic neurons in the intermediate and parvicellular reticular nuclei (IRt/PCRt) of the medulla oblongata, which are activated by NPY-triggered neural signaling from the hypothalamus, potentially through the nucleus tractus solitarius, mediate the NPY-induced inhibition of metabolic thermogenesis in brown adipose tissue (BAT) via their innervation of BAT sympathetic premotor neurons. Intriguingly, the GABAergic IRt/PCRt neurons innervating the BAT sympathetic premotor region also innervate the masticatory motor region, and stimulation of the IRt/PCRt elicits mastication and increases feeding as well as inhibits BAT thermogenesis. These results indicate that GABAergic IRt/PCRt neurons mediate hypothalamus-derived hunger signaling by coordinating both autonomic and feeding motor systems to reduce energy expenditure and to promote feeding.

Supratrigeminal Bilaterally Projecting Neurons Maintain Basal Tone and Enable Bilateral Phasic Activation of Jaw-Closing Muscles.

Anatomical studies have identified brainstem neurons that project bilaterally to left and right oromotor pools, which could potentially mediate bilateral muscle coordination. We use retrograde lentiviruses combined with a split-intein-mediated split-Cre-recombinase system in mice to isolate, characterize, and manipulate a population of neurons projecting to both the left and right jaw-closing trigeminal motoneurons. We find that these bilaterally projecting premotor neurons (BPNs) reside primarily in the supratrigeminal nucleus (SupV) and the parvicellular and intermediate reticular regions dorsal to the facial motor nucleus. These BPNs also project to multiple midbrain and brainstem targets implicated in orofacial sensorimotor control, and consist of a mix of glutamatergic, GABAergic, and glycinergic neurons, which can drive both excitatory and inhibitory inputs to trigeminal motoneurons when optogenetically activated in slice. Silencing BPNs with tetanus toxin light chain (TeNT) increases bilateral masseter activation during chewing, an effect driven by the expression of TeNT in SupV BPNs. Acute unilateral optogenetic inhibition of SupV BPNs identifies a group of tonically active neurons that function to lower masseter muscle tone, whereas unilateral optogenetic activation of SupV BPNs is sufficient to induce bilateral masseter activation both during resting state and during chewing. These results provide evidence for SupV BPNs in tonically modulating jaw-closing muscle tone and in mediating bilateral jaw closing. We developed a method that combines retrograde lentiviruses with the split-intein-split-Cre system in mice to isolate, characterize, and manipulate neurons that project to both left and right jaw-closing motoneurons. We show that these bilaterally projecting premotor neurons (BPNs) reside primarily in the supratrigeminal nucleus and the rostral parvicellular and intermediate reticular nuclei. BPNs consist of both excitatory and inhibitory populations, and also project to multiple brainstem nuclei implicated in orofacial sensorimotor control. Manipulation of the supratrigeminal BPNs during natural jaw-closing behavior reveals a dual role for these neurons in eliciting phasic muscle activation and in maintaining basal muscle tone. The retrograde lentivirus carrying the split-intein-split-Cre system can be applied to study any neurons with bifurcating axons innervating two brain regions.

Whole-brain mapping of afferent projections to the bed nucleus of the stria terminalis in tree shrews

The bed nucleus of the stria terminalis (BST) plays an important role in integrating and relaying input information to other brain regions in response to stress. The cytoarchitecture of the BST in tree shrews (Tupaia belangeri chinensis) has been comprehensively described in our previous publications. However, the inputs to the BST have not been described in previous reports. The aim of the present study was to investigate the sources of afferent projections to the BST throughout the brain of tree shrews using the retrograde tracer Fluoro-Gold (FG). The present results provide the first detailed whole-brain mapping of BST-projecting neurons in the tree shrew brain. The BST was densely innervated by the prefrontal cortex, entorhinal cortex, ventral subiculum, amygdala, ventral tegmental area, and parabrachial nucleus. Moreover, moderate projections to the BST originated from the medial preoptic area, supramammillary nucleus, paraventricular thalamic nucleus, pedunculopontine tegmental nucleus, dorsal raphe nucleus, locus coeruleus, and nucleus of the solitary tract. Afferent projections to the BST are identified in the ventral pallidum, nucleus of the diagonal band, ventral posteromedial thalamic nucleus, posterior complex of the thalamus, interfascicular nucleus, retrorubral field, rhabdoid nucleus, intermediate reticular nucleus, and parvicellular reticular nucleus. In addition, the different densities of BST-projecting neurons in various regions were analyzed in the tree shrew brains. In summary, whole-brain mapping of direct inputs to the BST is delineated in tree shrews. These brain circuits are implicated in the regulation of numerous physiological and behavioral processes including stress, reward, food intake, and arousal.

Injections of Algesic Solutions into Muscle Activate the Lateral Reticular Formation: A Nociceptive Relay of the Spinoreticulothalamic Tract.

Although musculoskeletal pain disorders are common clinically, the central processing of muscle pain is little understood. The present study reports on central neurons activated by injections of algesic solutions into the gastrocnemius muscle of the rat, and their subsequent localization by c-Fos immunohistochemistry in the spinal cord and brainstem. An injection (300μl) of an algesic solution (6% hypertonic saline, pH 4.0 acetate buffer, or 0.05% capsaicin) was made into the gastrocnemius muscle and the distribution of immunolabeled neurons compared to that obtained after control injections of phosphate buffered saline [pH 7.0]. Most labeled neurons in the spinal cord were found in laminae IV-V, VI, VII and X, comparing favorably with other studies, with fewer labeled neurons in laminae I and II. This finding is consistent with the diffuse pain perception due to noxious stimuli to muscles mediated by sensory fibers to deep spinal neurons as compared to more restricted pain localization during noxious stimuli to skin mediated by sensory fibers to superficial laminae. Numerous neurons were immunolabeled in the brainstem, predominantly in the lateral reticular formation (LRF). Labeled neurons were found bilaterally in the caudalmost ventrolateral medulla, where neurons responsive to noxious stimulation of cutaneous and visceral structures lie. Immunolabeled neurons in the LRF continued rostrally and dorsally along the intermediate reticular nucleus in the medulla, including the subnucleus reticularis dorsalis caudally and the parvicellular reticular nucleus more rostrally, and through the pons medial and lateral to the motor trigeminal nucleus, including the subcoerulear network. Immunolabeled neurons, many of them catecholaminergic, were found bilaterally in the nucleus tractus solitarii, the gracile nucleus, the A1 area, the CVLM and RVLM, the superior salivatory nucleus, the nucleus locus coeruleus, the A5 area, and the nucleus raphe magnus in the pons. The external lateral and superior lateral subnuclei of the parabrachial nuclear complex were consistently labeled in experimental data, but they also were labeled in many control cases. The internal lateral subnucleus of the parabrachial complex was labeled moderately. Few immunolabeled neurons were found in the medial reticular formation, however, but the rostroventromedial medulla was labeled consistently. These data are discussed in terms of an interoceptive, multisynaptic spinoreticulothalamic path, with its large receptive fields and role in the motivational-affective components of pain perceptions.

Injection of WGA-Alexa 488 into the ipsilateral hemidiaphragm of acutely and chronically C2 hemisected rats reveals activity-dependent synaptic plasticity in the respiratory motor pathways

WGA-Alexa 488 is a fluorescent neuronal tracer that demonstrates transsynaptic transport in the central nervous system. The transsynaptic transport occurs over physiologically active synaptic connections rather than less active or silent connections. Immediately following C2 spinal cord hemisection (C2Hx), when WGA-Alexa 488 is injected into the ipsilateral hemidiaphragm, the tracer diffuses across the midline of the diaphragm and retrogradely labels the phrenic nuclei (PN) bilaterally in the spinal cord. Subsequently, the tracer is transsynaptically transported bilaterally to the rostral Ventral Respiratory Groups (rVRGs) in the medulla over physiologically active connections. No other neurons are labeled in the acute C2Hx model at the level of the phrenic nuclei or in the medulla. However, with a recovery period of at least 7weeks (chronic C2Hx), the pattern of WGA-Alexa 488 labeling is notably changed. In addition to the bilateral PN and rVRG labeling, the chronic C2Hx model reveals fluorescence in the ipsilateral ventral and dorsal spinocerebellar tracts, and the ipsilateral reticulospinal tract. Furthermore, interneurons are labeled bilaterally in laminae VII and VIII of the spinal cord as well as neurons in the motor nuclei bilaterally of the intercostal and forelimb muscles. Moreover, in the chronic C2Hx model, there is bilateral labeling of additional medullary centers including raphe, hypoglossal, spinal trigeminal, parvicellular reticular, gigantocellular reticular, and intermediate reticular nuclei. The selective WGA-Alexa 488 labeling of additional locations in the chronic C2Hx model is presumably due to a hyperactive state of the synaptic pathways and nuclei previously shown to connect with the respiratory centers in a non-injured model. The present study suggests that hyperactivity not only occurs in neuronal centers and pathways caudal to spinal cord injury, but in supraspinal centers as well. The significance of such injury-induced plasticity is that hyperactivity may be a mechanism to re-establish lost function by compensatory routes which were initially physiologically inactive.

The pronociceptive dorsal reticular nucleus contains mostly tonic neurons and shows a high prevalence of spontaneous activity in block preparation

Despite the importance and significant clinical impact of understanding information processing in the nociceptive system, the functional properties of neurons in many parts of this system are still unknown. In this work we performed whole cell patch-clamp recording in rat brain stem blocks to characterize the electrophysiological properties of neurons in the dorsal reticular nucleus (DRt), a region known to be involved in pronociceptive modulation. We also compared properties of DRt neurons with those in the adjacent parvicellular reticular nucleus and in neighboring regions outside the reticular formation. We found that neurons in the DRt and parvicellular reticular nucleus had similar electrophysiological properties and exhibited mostly toniclike firing patterns, whereas neurons outside the reticular formation showed a larger diversity of firing patterns. Interestingly, more than one-half of the neurons also showed spontaneous activity. While the general view of the reticular formation, being a loosely associated mesh of groups of neurons with diverse function, and earlier reports suggests more electrophysiological heterogeneity, we showed that this is indeed not the case. Our results indicate that functional difference of neurons in the reticular formation may mostly be determined by their connectivity profiles and not by their intrinsic electrophysiological properties. The dominance of tonic neurons in the DRt supports previous conclusions that these neurons encode stimulus intensity through their firing frequency, while the high prevalence of spontaneous activity most likely shapes nociceptive modulation by this brain stem region.

Segregated anatomical input to sub-regions of the rodent superior colliculus associated with approach and defense.

The superior colliculus (SC) is responsible for sensorimotor transformations required to direct gaze toward or away from unexpected, biologically salient events. Significant changes in the external world are signaled to SC through primary multisensory afferents, spatially organized according to a retinotopic topography. For animals, where an unexpected event could indicate the presence of either predator or prey, early decisions to approach or avoid are particularly important. Rodents’ ecology dictates predators are most often detected initially as movements in upper visual field (mapped in medial SC), while appetitive stimuli are normally found in lower visual field (mapped in lateral SC). Our purpose was to exploit this functional segregation to reveal neural sites that can bias or modulate initial approach or avoidance responses. Small injections of Fluoro-Gold were made into medial or lateral sub-regions of intermediate and deep layers of SC (SCm/SCl). A remarkable segregation of input to these two functionally defined areas was found. (i) There were structures that projected only to SCm (e.g., specific cortical areas, lateral geniculate and suprageniculate thalamic nuclei, ventromedial and premammillary hypothalamic nuclei, and several brainstem areas) or SCl (e.g., primary somatosensory cortex representing upper body parts and vibrissae and parvicellular reticular nucleus in the brainstem). (ii) Other structures projected to both SCm and SCl but from topographically segregated populations of neurons (e.g., zona incerta and substantia nigra pars reticulata). (iii) There were a few brainstem areas in which retrogradely labeled neurons were spatially overlapping (e.g., pedunculopontine nucleus and locus coeruleus). These results indicate significantly more structures across the rat neuraxis are in a position to modulate defense responses evoked from SCm, and that neural mechanisms modulating SC-mediated defense or appetitive behavior are almost entirely segregated.

Muscle tone regulation during REM sleep: neural circuitry and clinical significance.

Rapid eye movement (REM) sleep is a distinct behavioral state characterized by an activated cortical and hippocampal electroencephalogram (EEG) and concurrent muscle atonia. Research conducted over the past 50 years has revealed the neuronal circuits responsible for the generation and maintenance of REM sleep, as well as the pathways involved in generating the cardinal signs of REM sleep such as cortical activation and muscle atonia. The generation and maintenance of REM sleep appear to involve a widespread network in the pons and medulla. The caudal laterodorsal tegmental nucleus (cLDT) and sublaterodorsal nucleus (SLD) within the dorsolateral pons contain REM-on neurons, and the ventrolateral periaqueductal grey (vlPAG) contains REM-off neurons. The interaction between these structures is proposed to regulate REM sleep amounts. The cLDT-SLD neurons project to the basal forebrain via the parabrachial-precoeruleus (PB-PC) complex, and this pathway may be critical for the EEG activation seen during REM sleep. Descending SLD glutamatergic projections activate the ventromedial medulla, and spinal cord interneurons mediate muscle atonia and suppress phasic muscle twitches in spinal musculature. In contrast, phasic muscle twitches in the masseter muscles may be driven by glutamatergic neurons in the rostral parvicellular reticular nucleus (PCRt); however, the brain region responsible for generating phasic twitches in the other cranial muscles including facial muscles and tongue are not clear.

Retrograde Neuron Tracing with Microspheres Reveals Projection of CGRP-Immunolabeled Vestibular Afferent Neurons to the Vestibular Efferent Nucleus in the Brainstem of Rats

Objective: A new retrograde neuron-tracing technique with microspheres was used to explore the possible innervation of calcitonin gene-related peptide (CGRP)-immunolabeled vestibular afferent neurons in the vestibular efferent immunolabeled nucleus in the brainstem. Methods: 0.1 µl of 5% microfluorospheres was injected into the area of the vestibular efferent nucleus, which is located lateral to the genu of the facial nerve. CGRP immunohistochemistry was processed in serial sections of the brainstem at the facial nerve genu level. Double-labeled neurons with both CGRP immunoreactivity and microfluorospheres were examined with fluorescence and confocal laser microscopy. Results: Three types of labeled neurons were observed: (1) neurons only retrogradely microfluorosphere-labeled that were mainly located in the medial vestibular nucleus, lateral vestibular nucleus, superior vestibular nucleus and parvicellular reticular nucleus on the ipsilateral side of the injection; (2) neurons that were both immunolabeled with CGRP and also retrogradedly labeled with microfluorospheres, indicating that they are CGRP cells projecting to the area of vestibular efferent nucleus, these cells were mainly distributed in the superior vestibular nucleus and dorsal vestibular nucleus, and (3) cells only immunolabeled for CGRP that were scattered extensively in the brainstem. Conclusion: The presented methodical contribution demonstrates the suitability of fluorescein-labeled microspheres for retrograde neuronal tracing. The vestibular nuclei contain numerous afferent neurons that send projections to the vestibular efferent nucleus, some of which are CGRP cells. This afferent innervation provides morphological evidence that the vestibular efferent neurons receive input from the vestibular afferent neurons including CGRP cells. These vestibular primary CGRP afferent neurons may have an influence on vestibular efferent neurons. CGRP acts as an important co-transmitter or modulator in the afferent-mediated activity of vestibular efferent neurons, which in turn affect afferents in the vestibular end organs.

Migratory routes and fates of cells transcribing the Wnt‐1 gene in the murine hindbrain

To investigate the origins, migrations, and fates of Wnt-1-expressing cells in the murine hindbrain, mice carrying a Wnt-1 enhancer/lacZ transgene were observed from embryonic day (E) 8 through postnatal day 18. The transgene-stained ventricular layer waxed and waned prior to and following migrations from it. Stained cells migrated first external to the hindbrain as neural crest and then within it to form typical populations of the rhombic lip, as well as others not recognized as lip derivatives. Migrations originated in a temporally defined sequence, many from discrete rhombomeres. All moved first radially, then rostrally and/or ventrally, ipsi-, or contralaterally, in the mantle or marginal layers. These movements ultimately formed elements of several nuclei, aligned in four longitudinal bands: dorsal (including the gracile, cuneate, cochlear, and vestibular nuclei, plus cerebellar granular cells), dorsal intermediate (including trigeminal sensory, parvicellular reticular, and deep cerebellar nuclei), ventral intermediate (including lateral and intermediate reticular nuclei), and ventral (including the raphe obscurus and pontine nuclei). Transgene staining often persisted long enough to identify stained cells in their definitive, adult nuclei. However, staining was transient. The strength of the staining, however, was in its ability to reveal origins and migrations in both whole-mounts and sections, in single cell detail. The present results will permit analyses of the effects of genetic manipulations on Wnt-1 lineage cells.

The rat ponto-medullary network responsible for paradoxical sleep onset and maintenance: a combined microinjection and functional neuroanatomical study.

The neuronal network responsible for paradoxical sleep (PS) onset and maintenance has not previously been identified in the rat, unlike the cat. To fill this gap, this study has developed a new technique involving the recording of sleep-wake states in unanaesthetized head-restrained rats whilst locally administering pharmacological agents by microiontophoresis from glass multibarrel micropipettes, into the dorsal pontine tegmentum and combining this with functional neuroanatomy. Pharmacological agents used for iontophoretic administration included carbachol, kainic acid, bicuculline and gabazine. The injection sites and their efferents were then identified by injections of anterograde (phaseolus vulgaris leucoagglutinin) or retrograde (cholera toxin B subunit) tracers through an adjacent barrel of the micropipette assembly and by C-Fos immunostaining. Bicuculline, gabazine and kainic acid ejections specifically into the pontine sublaterodorsal nucleus (SLD) induced within a few minutes a PS-like state characterized by a continuous muscle atonia, low voltage EEG and a lack of reaction to stimuli. In contrast, carbachol ejections into the SLD induced wakefulness. In PHA-L, glycine and C-Fos multiple double-labelling experiments, anterogradely labelled fibres originating from the SLD were seen apposed on glycine and C-Fos positive neurons (labelled after 90 min of pharmacologically induced PS-like state) from the ventral gigantocellular and parvicellular reticular nuclei. Altogether, these data indicate that the SLD nuclei contain a population of neurons playing a crucial role in PS onset and maintenance. Furthermore, they suggest that GABAergic disinhibition and glutamate excitation of these neurons might also play a crucial role in the onset of PS.

Brainstem projections to midline and intralaminar thalamic nuclei of the rat.

The projections from the brainstem to the midline and intralaminar thalamic nuclei were examined in the rat. Stereotaxic injections of the retrograde tracer cholera toxin beta -subunit (CTb) were made in each of the intralaminar nuclei of the dorsal thalamus: the lateral parafascicular, medial parafascicular, central lateral, paracentral, oval paracentral, and central medial nuclei; in the midline thalamic nuclei-the paraventricular, intermediodorsal, mediodorsal, paratenial, rhomboid, reuniens, and submedius nuclei; and, in the anteroventral, parvicellular part of the ventral posterior, and caudal ventral medial nuclei. The retrograde cell body labeling pattern within the brainstem nuclei was then analyzed. Nearly every thalamic site received a projection from the deep mesencephalic reticular, pedunculopontine tegmental, dorsal raphe, median raphe, laterodorsal tegmental, and locus coeruleus nuclei. Most intralaminar thalamic sites were also innervated by unique combinations of medullary and pontine reticular formation nuclei such as the subnucleus reticularis dorsalis, gigantocellular, dorsal paragigantocellular, lateral, parvicellular, caudal pontine, ventral pontine, and oral pontine reticular nuclei; the dorsomedial tegmental, subpeduncular tegmental, and ventral tegmental areas; and, the central tegmental field. In addition, most intralaminar injections resulted in retrograde cell body labeling in the substantia nigra, nucleus Darkschewitsch, interstitial nucleus of Cajal, and cuneiform nucleus. Details concerning the pathways from the spinal trigeminal, nucleus tractus solitarius, raphe magnus, raphe pallidus, and the rostral and caudal linear raphe nuclei to subsets of midline and intralaminar thalamic sites are discussed in the text. The discussion focuses on brainstem-thalamic pathways that are likely involved in arousal, somatosensory, and visceral functions.