Olfactory bulbectomy reduces cerebral glucose utilization: 2-[14C]deoxyglucose autoradiographic study

The olfactory bulbectomized (OBX) rat is an extensively investigated animal model of depression. In the present study the effects of olfactory bulbectomy in drug-naive adult male Sprague–Dawley rats (200–240 g) on global (gCGU) and regional cerebral glucose (rCGU) utilization was evaluated. Two weeks following surgery, the autoradiographic measurement of CGU using [¹⁴C]-2-deoxyglucose was employed. The levels of CGU in the OBX and sham-operated rats were compared in 40 brain regions. Statistical methods indicate significantly lower levels of global (overall) CGU in the OBX group than in the sham group. Discriminant analysis was done on the z-scores to remove animal to animal variability. The following thirteen regions were identified by the stepwise discriminant analysis of the z-scores as significantly contributing to the differences between the sham and OBX: amygdala, cingulate cortex, caudate putamen at the level of globus pallidus, caudate putamen-lateral part, dorsal subiculum, dorsal thalamus, hypothalamus, median raphe, somatosensory cortex, substantia nigra, ventral hippocampus, ventral tegmental area and the ventral thalamus. The pattern of changes in the rCGU following OBX does not completely correlate with the pattern of connectivity of the olfactory bulbs, however, many regions with direct connection to the olfactory bulbs (e.g., amygdala, hypothalamus, ventral hippocampus, and ventral tegmental area) were found to be important for differentiation. No left to right asymmetries in the rCGU were found. The data suggest that there are very important regional differences in glucose utilization between the OBX and sham operated rats, which points to the need to study antidepressants in an animal model of depression rather than in normal animals.     

Absence of μ opioid receptor mRNA expression in astrocytes and microglia of rat spinal cord

Cumulating evidence has demonstrated that μ opioid receptor (MOR) agonists promote spinal glial activation, lead to synthesis and release of proinflammatory cytokines and chemokines, and contribute to opioid-induced hyperalgesia and development of opioid tolerance and dependence. However, whether these MOR agonists directly or indirectly act on spinal cord astrocytes and microglial cells in vivo is unclear. In the present study, by combining the techniques of in-situ hybridization of MOR mRNA with immunohistochemistry of glial fibrillary acidic protein (GFAP; an astrocyte marker) and Iba1 (a microglial marker), we examined expression and distribution of GFAP, Iba1, and MOR mRNA in the spinal cord of rats under chronic morphine tolerance conditions. Intrathecal injections of morphine twice daily for 7 days reduced morphine analgesic effect and increased both GFAP and Iba1 immunostaining densities in the spinal cord. Surprisingly, neither GFAP nor Iba1 colocalized with MOR mRNA in spinal cord cells. Our findings indicate that MOR expression is absent from spinal cord astrocytes and microglia, suggesting that these cell types are indirectly activated by MOR agonists under chronic opioid tolerance conditions.     

Morphology and axonal projection pattern of neurons in the telencephalon of the fire-bellied toad Bombina orientalis: an anterograde, retrograde, and intracellular biocytin labeling study

The connectivity and cytoarchitecture of telencephalic centers except dorsal and medial pallium were studied in the fire-bellied toad Bombina orientalis by anterograde and retrograde biocytin labeling and intracellular biocytin injection (total of 148 intracellularly labeled neurons or neuron clusters). Our findings suggest the following telencephalic divisions: (1) a central amygdala-bed nucleus of the stria terminalis in the caudal midventral telencephalon, connected to visceral-autonomic centers; (2) a vomeronasal amygdala in the caudolateral ventral telencephalon receiving input from the accessory olfactory bulb and projecting mainly to the preoptic region/hypothalamus; (3) an olfactory amygdala in the caudal pole of the telencephalon lateral to the vomeronasal amygdala receiving input from the main olfactory bulb and projecting to the hypothalamus; (4) a medial amygdala receiving input from the anterior dorsal thalamus and projecting to the medial pallium, septum, and hypothalamus; (5) a ventromedial column formed by a nucleus accumbens and a ventral pallidum projecting to the central amygdala, hypothalamus, and posterior tubercle; (6) a lateral column constituting the dorsal striatum proper rostrally and the dorsal pallidum caudally, and a ventrolateral column constituting the ventral striatum. We conclude that the caudal mediolateral complex consisting of the extended central, vomeronasal, and olfactory amygdala of anurans represents the ancestral condition of the amygdaloid complex. During the evolution of the mammalian telencephalon this complex was shifted medially and involuted. The mammalian basolateral amygdala apparently is an evolutionary new structure, but the medial portion of the amygdalar complex of anurans reveals similarities in input and output with this structure and may serve similar functions.     

Distribution of glial fibrillary acidic protein-immunopositive structures in the brain of the domestic chicken (Gallus domesticus)

The present study is the first comprehensive mapping of glial fibrillary acidic protein (GFAP)-immunopositive structures in the avian brain. Two main types of GFAP-immunopositive elements were observed: (1) nonbranching fibers, occasionally twisted or varicose, and (2) star-shaped cells. Long immunostained fibers orignate from the lateral ventricle to form three bundles. Fibers of the dorsal group, emanating from the dorsal/lateral corner of the ventricle, course in lateral, anterior, and ventral directions forming a semidome, which separates the outer pallial (lateral cortical) regions from the underlying striatal mass. The middle group of fibers is directed anteriorly and laterally corresponding to the laminae frontales superior and suprema. The ventral fiber bundle is conical and traverses the lobus parolfactorius, crossing also the lamina medullaris dorsalis (the latter consisting mainly of star-shaped cells). The hippocampus, septum, and hypothalamus also contain straight radial fibers. In some areas, given their variable orientation, the fibers cannot be regarded as merely persisting radial glia. In the telencephalon, the nuclei basalis, accumbens, ectostriatum, paleostriatum primitivum, and the ventral paleostriatum are particularly rich in GFAP-positive cells, whereas the neostriatum, hyperstriatum, and paleostriatum augmentatum are almost devoid of GFAP labelling. Certain nuclei of the thalamus and the lower brainstem are conspicuous by their low levels of GFAP immunoreactivity. The Bergmann glia were GFAP-immunonegative.     

Efferent projections of reuniens and rhomboid nuclei of the thalamus in the rat

The nucleus reuniens (RE) is the largest of the midline nuclei of the thalamus and exerts strong excitatory actions on the hippocampus and medial prefrontal cortex. Although RE projections to the hippocampus have been well documented, no study using modern tracers has examined the totality of RE projections. With the anterograde anatomical tracer Phaseolus vulgaris leuccoagglutinin, we examined the efferent projections of RE as well as those of the rhomboid nucleus (RH) located dorsal to RE. Control injections were made in the central medial nucleus (CEM) of the thalamus. We showed that the output of RE is almost entirely directed to the hippocampus and "limbic" cortical structures. Specifically, RE projects strongly to the medial frontal polar, anterior piriform, medial and ventral orbital, anterior cingulate, prelimbic, infralimbic, insular, perirhinal, and entorhinal cortices as well as to CA1, dorsal and ventral subiculum, and parasubiculum of the hippocampus. RH distributes more widely than RE, that is, to several RE targets but also significantly to regions of motor, somatosensory, posterior parietal, retrosplenial, temporal, and occipital cortices; to nucleus accumbens; and to the basolateral nucleus of amygdala. The ventral midline thalamus is positioned to exert significant control over fairly widespread regions of the cortex (limbic, sensory, motor), hippocampus, dorsal and ventral striatum, and basal nuclei of the amygdala, possibly to coordinate limbic and sensorimotor functions. We suggest that RE/RH may represent an important conduit in the exchange of information between subcortical-cortical and cortical-cortical limbic structures potentially involved in the selection of appropriate responses to specific and changing sets of environmental conditions.     

Alterations in the Expression of Genes That Encode Subunits of Ionotropic Glutamate Receptors and the Glutamate Transporter in Brain Structures of Rats after Psychogenic Stress

We studied modifications in the glutamatergic system of the brain as a factor in the development of post-traumatic stress disorder. An analysis of mRNA production of NMDA (GluN1, GluN2a, and GluN2b) and AMPA (GluA1 and GluA2) glutamate receptors, as well as the EAAT2 glutamate transporter was performed in the brain of rats subjected to stress associated with contact with a predator (a black-tailed python). Studies were performed in 6 or 24 h as well as in 3, 9, and 25 days after stress. The most-pronounced alterations of expression of all studied genes were revealed 25 days after stress. The level of EAAT2 mRNA increased in the ventral hippocampus. The expression of the genes that encode GluA1 and GluA2 subunits of AMPA receptors decreased in the dorsal and increased in the ventral hippocampus. The changes in the expression of the gene that encodes the GluN2b subunit of the NMDA receptor were also region specific. In the ventral hippocampus and medial prefrontal cortex we observed an increase in the expression of GluN2b mRNA, while it decreased in the dorsal hippocampus. The increased expression of the gene that encodes the GluN2a subunit was found in the amygdala. These alterations may be a mechanism of the development of delayed post-stress neurological–psychiatric impairments.     

Serotonin neurons of the midbrain raphe: ascending projections.

The ascending projections of serotonin neurons of the midbrain raphe were analyzed in the rat using the autoradiographic tracing method. Axons of raphe serotonin neurons ascend in the ventral tegmental area and enter the medial forebrain bundle. A number of fibers leave the major group to ascend along the fasciculus retroflexus. Some fibers enter the habenula but the majority turn rostrally in the internal medullary lamina of the thalamus to innervate dorsal thalamus. Two additional large projections leave the medial forebrain bundle in the hypothalamus; the ansa peduncularis-ventral amygdaloid bundle system turns laterally through the internal capsule into the striatal complex, amygdala and the external capsule to reach lateral and posterior cortex, and another system of fibers turns medially to innervate medial hypothalamus and median eminence and form a contrelateral projection via the supraoptic commissures. Rostrally the major group in the medial forebrain bundle divides into several components: fibers entering the stria medullaris to terminate in thalamus; fibers entering the stria terminalis to terminate in the amygdala; fibers traversing the fornix to the hippocampus; fibers running through septum to enter the cingulum and terminate in dorsal and medial cortex and in hippocampus; fibers entering the external capsule to innervate rostral and lateral cortex; and fibers continuing forward in the medial olfactory stria to terminate in the anterior olfactory nucleus and olfactory bulb.     

Induction of immunoreactive interleukin-1β and tumor necrosis factor-α in the brains of rabies virus infected rats

Interleukin-1β (IL-1β) and tumor necrosis factor-α (TNFα) are important cytokines in the development of brain inflammation during pathological process. During rabies virus infection, the level of these proinflammatory cytokines are enhanced in the brain. In the present study we determined the cellular localization of these two cytokines by immunocytochemistry in brains of rats infected with rabies virus, at different time-intervals of the disease (day 1, 3, 4, 5 and at final stage day 6 post-infection (p.i.)). Cellular identification of IL-1β (irIL-1β) and TNFα (irTNFα) immunopositive cells was studied using a polyclonal antibody against these cytokines and against glial fibrillary acidic protein (GFAP) to detect astrocytes and GSA-I-B4 isolectin to detect microglial cells and/or infiltrating macrophages. In brains of control and early infected rats, irIL-1β was only detected in fibers located in the hypothalamus, supraoptic and tractus optic nuclei and infundibular nucleus. From day 4 onwards until day 6 p.i., enhanced irIL-1β was found and identified either in activated ameboid and/or infiltrated macrophages (amygdala, thalamus, internal capsula, subtantia nigra, septal nuclei and around blood vessels), or in activated ramified cells (hypothalamus and periventricular nucleus, piriformis and cingulate cortex, hippocampus). IrTNFα was observed in the brains of rats at a final stage of disease (day 5 and 6 p.i.): in the hypothalamus, the amygdala, the internal capsula, the thalamus, the septal nuclei, the hippocampus, the habenular nuclei and around the blood vessels. Ir-TNFα was detected in round cells identified as ameboid microglia and/or infiltrated macrophages. A marked activation of microglial and astroglial cells was observed mainly in the hypothalamus, the thalamus and hippocampus and around the blood vessels, at day 4 p.i. and later, revealing a high central inflammatory reaction in brains of rabies virus infected rats. These results showed that IL-1β and TNFα are produced in the brain both by local microglial cells and infiltrating macrophages during rabies infection. Thus, these cytokines may play an important role in coordinating the dramatic inflammatory response associated with the rabies-encephalopathy as well as in the neural modification and alteration of brain functions.     

Localization of B1 bradykinin receptor mRNA in the primate brain and spinal cord: an in situ hybridization study

The bradykinin 1 and 2 receptors (B1R, B2R) are important mediators of cardiovascular homeostasis, inflammation, and nociception. While B2R is constitutively expressed in many tissues, B1R expression is thought to be absent, but induced under proinflammatory conditions. However, recent data from knockout mice have indicated that B1R acts centrally to mediate nociception, a finding that suggests the constitutive presence of B1R in brain and/or spinal cord. The purpose of the present study was to further elucidate the physiological role of B1R by evaluating the localization of B1R mRNA in the nonhuman primate brain and spinal cord with in situ hybridization. Cryostat sections from monkey brain and spinal cord were hybridized with a [(35)S]-labeled riboprobe complementary to B1R mRNA, stringently washed, and apposed to film and emulsion. The results of these studies revealed the presence of B1R mRNA throughout the rostral-caudal extent of the brain and spinal cord. In particular, labeled cells were seen in the cerebral and entorhinal cortex, dentate gyrus, and pyramidal neurons of the hippocampus, in the thalamus, hypothalamus, amygdala, pontine nuclei, spinal cord, and dorsal root ganglion. Together the present findings offer detailed information about the distribution of B1R mRNA in the primate brain and spinal cord and demonstrate a basal level of expression in the primate nervous system. Moreover, these data provide a foundation for understanding the central actions of kinins and their putative role in mediating a number of processes, including pain and nociception.     

A meta‐analysis of deep brain structural shape and asymmetry abnormalities in 2,833 individuals with schizophrenia compared with 3,929 healthy volunteers via the ENIGMA Consortium

Schizophrenia is associated with widespread alterations in subcortical brain structure. While analytic methods have enabled more detailed morphometric characterization, findings are often equivocal. In this meta‐analysis, we employed the harmonized ENIGMA shape analysis protocols to collaboratively investigate subcortical brain structure shape differences between individuals with schizophrenia and healthy control participants. The study analyzed data from 2,833 individuals with schizophrenia and 3,929 healthy control participants contributed by 21 worldwide research groups participating in the ENIGMA Schizophrenia Working Group. Harmonized shape analysis protocols were applied to each site''s data independently for bilateral hippocampus, amygdala, caudate, accumbens, putamen, pallidum, and thalamus obtained from T1‐weighted structural MRI scans. Mass univariate meta‐analyses revealed more‐concave‐than‐convex shape differences in the hippocampus, amygdala, accumbens, and thalamus in individuals with schizophrenia compared with control participants, more‐convex‐than‐concave shape differences in the putamen and pallidum, and both concave and convex shape differences in the caudate. Patterns of exaggerated asymmetry were observed across the hippocampus, amygdala, and thalamus in individuals with schizophrenia compared to control participants, while diminished asymmetry encompassed ventral striatum and ventral and dorsal thalamus. Our analyses also revealed that higher chlorpromazine dose equivalents and increased positive symptom levels were associated with patterns of contiguous convex shape differences across multiple subcortical structures. Findings from our shape meta‐analysis suggest that common neurobiological mechanisms may contribute to gray matter reduction across multiple subcortical regions, thus enhancing our understanding of the nature of network disorganization in schizophrenia.     

Estrogen decreases levels of calcineurin in rat amygdala and hippocampus

We examined the effects of estradiol benzoate (E2) on the protein expression of calcineurin in amygdaloid and hippocampal structures of ovariectomized (OVX) rats. Significant decreases in levels of calcineurin immunolabeling were seen in the medial and basomedial, but not central or basolateral, amygdala. Estrogen also reduced calcineurin immunoreactivity in the CA1 region of the hippocampus, but not in the CA3 region, hilus or ventral or dorsal dentate gyrus structures of hippocampus. These results indicate that E2 acts on calcineurin in a neuroanatomically specific manner and may be involved in estrogen-mediated regulation of gene expression.     

Effects of a selective Y2R antagonist, JNJ-31020028, on nicotine abstinence-related social anxiety-like behavior, neuropeptide Y and corticotropin releasing factor mRNA levels in the novelty-seeking phenotype

An outbred rat model of novelty-seeking phenotype has predictive value for the expression of locomotor sensitization to nicotine. When experimentally naïve rats are exposed to a novel environment, some display high rates of locomotor reactivity (HRs, scores ranking at top 1/3rd of the population), whereas some display low rates (LRs, scores ranking at bottom 1/3rd of the population). Basally, HRs display lower anxiety-like behavior compared to LRs along with higher neuropeptide Y (NPY) mRNA in the amygdala and the hippocampus. Following an intermittent behavioral sensitization to nicotine regimen and 1 wk of abstinence, HRs show increased social anxiety-like behavior in the social interaction test and robust expression of locomotor sensitization to a low dose nicotine challenge. These effects are accompanied by a deficit in NPY mRNA levels in the medial nucleus of the amygdala and the CA3 field of the hippocampus, and increases in Y2R mRNA levels in the CA3 field and corticotropin releasing factor (CRF) mRNA levels in the central nucleus of the amygdala. Systemic and daily injections of a Y2R antagonist, JNJ-31020028, during abstinence fully reverse nicotine-induced social anxiety-like behavior, the expression of locomotor sensitization to nicotine challenge, the deficit in the NPY mRNA levels in the amygdala and the hippocampus, as well as result an increase in Y2R mRNA levels in the hippocampus and the CRF mRNA levels in the amygdala in HRs. These findings implicate central Y2R in neuropeptidergic regulation of social anxiety in a behavioral sensitization to nicotine regimen in the LRHR rats.     

The Effect of Minocycline on Motor Neuron Recovery and Neuropathic Pain in a Rat Model of Spinal Cord Injury

Objective

Minocycline, a second-generation tetracycline-class antibiotic, has been well established to exert a neuroprotective effect in animal models and neurodegenerative disease through the inhibition of microglia. Here, we investigated the effects of minocycline on motor recovery and neuropathic pain in a rat model of spinal cord injury.

Methods

To simulate spinal cord injury, the rats'' spinal cords were hemisected at the 10th thoracic level (T10). Minocycline was injected intraperitoneally, and was administered 30 minutes prior surgery and every second postoperative day until sacrifice 28 days after surgery. Motor recovery was assessed via the Basso-Beattie-Bresnahan test. Mechanical hyperalgesia was measured throughout the 28-day post-operative course via the von Frey test. Microglial and astrocyte activation was assessed by immunohistochemical staining for ionized calcium binding adaptor molecule 1 (Iba1) and glial fibrillary acidic protein (GFAP) at two sites: at the level of hemisection and at the 5th lumbar level (L5).

Results

In rats, spinal cord hemisection reduced locomotor function and induced a mechanical hyperalgesia of the ipsilateral hind limb. The expression of Iba1 and GFAP was also increased in the dorsal and ventral horns of the spinal cord at the site of hemisection and at the L5 level. Intraperitoneal injection of minocycline facilitated overall motor recovery and attenuated mechanical hyperalgesia. The expression of Iba1 and GFAP in the spinal cord was also reduced in rats treated with minocycline.

Conclusion

By inhibiting microglia and astrocyte activation, minocycline may facilitate motor recovery and attenuate mechanical hyperalgesia in individuals with spinal cord injuries.     

Cerebral uptake of [14C]deoxyglucose during the entire seizure and the recovery period in an El mouse

Local cerebral glucose utilization (LCGU) was examined in an El mouse during an entire seizure (tonic-clonic convulsions and recovery), and during the recovery period only. LCGU was measured quantitatively in 18 structures. In the whole-seizure group, the parietal cortex, dorsal hippocampus, dentate gyrus, ventral thalamus and cerebellar nuclei showed a significant increase in the uptake of 2-DG. In the recovery-period group, compared with the control group, a relative increase was found in the frontal, temporal and occipital cortex, amygdala, substantia grisea centralis mesencephali, cerebellar nuclei and caudate putamen, as well as the parietal cortex, dorsal and ventral hippocampus and dentate gyrus. To summarize, the hyperactivity in the parietal cortex, dorsal hippocampus, dentate gyrus, amygdala and cerebellar nuclei continued throughout the convulsive and recovery periods.     

Morphology,axonal projection pattern,and responses to optic nerve stimulation of thalamic neurons in the fire-bellied toad Bombina orientalis

Intracellular recording and biocytin labeling were carried out in the fire-bellied toad Bombina orientalis to study the morphology and axonal projections of thalamic (TH) neurons and their responses to electrical optic nerve stimulation. Labeled neurons (n = 142) were divided into the following groups: TH1 neurons projecting to the dorsal striatum; TH2 neurons projecting to the amygdala, nucleus accumbens, and septal nuclei; TH3 neurons projecting to the medial or dorsal pallium; TH4 neurons with projections ascending to the dorsal striatum or ventral striatum/amygdala and descending to the optic tectum, tegmentum, and rostral medulla oblongata; TH5 neurons with projections to the tegmentum, rostral medulla oblongata, prectectum, or tectum; and TH6 neurons projecting to the hypothalamus. TH1 neurons are found in the central, TH2 neurons in the anterior and central, TH3 neurons in the anterior dorsal nucleus, and TH4 and TH5 neurons in the posterior dorsal or ventral nucleus. Neurons with descending projections arborize in restricted parts of retinal afferents; neurons with ascending projections do not substantially arborize within retinal afferents. At electrical optic nerve stimulation, neurons in the ventral thalamus respond with excitation at latencies of 10.8 msec; one-third of them follow repetitive stimulation and possibly are monosynaptically driven. Neurons in the dorsal thalamus respond mostly with inhibition at latencies of 42.3 msec and are polysynaptically driven. This corroborates the view that neurons in the dorsal thalamus projecting to the telencephalon receive no substantial direct retinal input and that the thalamopallial pathway of amphibians is not homologous to the mammalian retinogeniculocortical pathway.     

Widespread expression in adult rat forebrain of mRNA encoding high-affinity neurotensin receptor

The peptides neurotensin (NT) and neuromedin N exert effects on neurons by means of a high-affinity NT receptor (NTRH) belonging to the superfamily of G-protein–coupled receptors. In the present study, we used in situ hybridization histochemistry with sensitive riboprobe methodology to investigate the distribution of NTRH mRNA in the forebrain of adult rats. Labeled cells were abundant in the hypothalamus, epithalamus, ventral thalamus, septum, amygdala, and pallidum, including many regions where NTRH mRNA had not been detected previously. In the hypothalamus, novel sites of NTRH mRNA expression included the arcuate, periventricular, paraventricular, supraoptic, medial preoptic, anterior, ventromedial, and posterior nuclei, as well as the lateral hypothalamic area. In the thalamus, novel sites of expression included the anterodorsal nucleus, lateral habenula, and zona incerta, where labeling was much more extensive than previously reported. Novel telencephalic sites of expression included most bed nuclei of the stria terminalis, most divisions of the amygdala, the main olfactory bulb, the endopiriform nucleus, the claustrum, many parts of retrohippocampal allocortex, and limited parts of most isocortical areas. Novel sites of expression were also observed in the midbrain and pons. Taking into account expected differences in the subcellular locations of receptor mRNA and protein, the regional distribution of NTRH mRNA agrees well with that of NTRH determined previously. Our results identify many novel sites of NTRH mRNA expression in adult brain and provide a basis for investigating involvement of NT and related peptides in regulating the activity of these diverse cells, whose phenotypes remain largely undetermined. J. Comp. Neurol. 402:475–500, 1998. Published 1998 Wiley-Liss, Inc.