Repeated social defeat stress exacerbates lipopolysaccharide-induced behavioural deficits in mice: ameliorative role of Chrysophyllum albidum fruit extract through anti-neuroinflammation, antioxidant and neurochemical balance.

Development of neuropsychiatric disorder is associated with stress-related increase in pro-inflammatory cytokines. Chrysophyllum albidum fruit is an edible tropical fruit containing vitamins and phenolic compounds, well known for their anti-inflammatory and antioxidant activities. This study was designed to investigate the neuroprotective effect of C. albidum fruit extract (CAFE) on stress and lipopolysaccharide (LPS)-induced behavioral and neurochemical impairments in mice. Male Swiss mice were divided into 6 groups (n = 6). Groups 1-3 were orally treated daily for 14 days with normal saline (0.1 mL/10 g), CAFE (100 mg/kg) and Ferulic acid (FA, 10 mg/kg), and left in home cage as controls. Groups 4-6 were treated similarly but subjected to repeated social defeat (RSD) stress using the resident-intruder model from days 1-14. The RSD-animals were injected with LPS (125 µg/kg, i.p) 60 min after each RSD session from days 8-14. Neurobehavioral functions: locomotor, cognitive and anxiety-like behaviors were assessed 24 h after the last treatment. Pro-inflammatory cytokines (IL-1ß, IL-6 and TNF-α), dopamine, acetylcholinesterase, glutamic acid decarboxylase (GAD), malondialdehyde, nitrites, and reduced glutathione (GSH) were determined in brain tissue. CAFE significantly attenuated RSD and LPS-induced hypolocomotion, cognitive impairment and anxiety-like behavior when compared to the control. Treatment with CAFE also significantly reversed the negative effects of RSD and LPS on pro-inflammatory cytokines, dopamine, acetylcholinesterase, GAD, and oxidative-nitrosative stress levels. The findings clearly indicated that Chrysophyllum albidum fruit demonstrated neuroprotective effects and can play a key role in mitigating against chronic stress and inflammation linked to neuropsychiatric disorders.

Placental tissue of greenhouse muskmelon (Cucumis melo L.) contains more gamma-aminobutyric acid with antioxidant capacity than the fleshed pulp.

Our previous study revealed that gamma-aminobutyric acid (GABA) in Earl's muskmelon is more concentrated in the inner than the outer parts of the fruit. Here, the GABA and antioxidant capacity of the placental tissue of muskmelon, which is considered waste, were evaluated for possible use as a source of bioactive compounds. The concentrations of GABA and related substances in the placental tissue were significantly higher than in the fleshed pulp, whereas glutamic acid and sugar levels were significantly lower. The two sites showed no difference in GAD activity. Furthermore, the placental site showed high antioxidant capacities based on 2,2-diphenyl-1-picrylhydrazyl and oxygen radical absorbance capacity for hydrophilic compounds assays compared with the fleshed pulp, because of the higher levels of total phenolic and L-ascorbic acids. Therefore, the placental tissue of muskmelons may be useful for developing functional foods, which would also reduce the amount of residues during muskmelon processing.

Development of a dual-fluorescence reporter system for high-throughput screening of L-aspartate-α-decarboxylase.

ß-Alanine (3-aminopropionic acid) holds great potential in industrial application. It can be obtained through a chemical synthesis route, which is hazardous to the environment. It is well known that l-aspartate-α-decarboxylase (ADC) can convert l-aspartate to ß-alanine in bacteria. However, due to the low activity of ADC, industrial production of ß-alanine through the green biological route remains unclear. Thus, improving the activity of ADC is critical to reduce the cost of ß-alanine production. In this study, we established a dual-fluorescence high-throughput system for efficient ADC screening. By measuring the amount of ß-alanine and the expression level of ADC using two different fluorescence markers, we can rapidly quantify the relative activity of ADC variants. From a mutagenesis library containing 2000 ADC variants, we obtained a mutant with 33% increased activity. Further analysis revealed that mutations of K43R and P103Q in ADC significantly improved the yield of ß-alanine produced by the whole-cell biocatalysis. Compared with the previous single-fluorescence method, our system can not only quantify the amount of ß-alanine but also measure the expression level of ADC with different fluorescence, making it able to effectively screen out ADC variants with improved relative activity. The dual-fluorescence high-throughput system for rapid screening of ADC provides a good strategy for industrial production of ß-alanine via the biological conversion route in the future.

The mechanism of freeze-thawing induced accumulation of γ-aminobutyric acid in germinated soybean.

BACKGROUND: γ-Aminobutyric acid (GABA) is a non-protein amino acid with several functions in the human body. Although freeze-thawing could effectively accumulate GABA in soybean sprouts, the mechanism has not been revealed. The mechanism by which freeze-thawing enhances GABA accumulation in germinated soybean was revealed by evaluating GABA content, the activity of related synthesis enzymes, and the microstructure of the tissues and cells of sprouts. The germinated soybeans were treated at different temperatures (from -196 °C to 25 °C) for 12 h and then thawed at 25 °C for 6 h. RESULTS: The results showed that GABA content in frozen soybean sprouts did not change significantly before thawing. After thawing, the GABA content of sprouts increased by 83.9% and 82.9% when treated by liquid nitrogen flash freeze at - 80 °C for 12 h compared with the control (4 °C treatment for 12 h). The results indicated that GABA formation mainly occurred during thawing. However, glutamate decarboxylase (GAD), diamine oxidase (DAO), and aminoaldehyde dehydrogenase (AMADH) activity decreased during thawing. Based on the malonaldehyde (MDA) content and microstructure of sprouts, it was suggested that freezing at lower temperatures (< -20 °C) maintained the integrity of the cell structure, while the tissues and cell membranes were broken during thawing. CONCLUSION: These results could provide evidence for the hypothesis that GABA formation resulted from full contact between enzymes and substrates during thawing, rather than the contribution of higher enzyme activity. © 2019 Society of Chemical Industry.

Estrogen Treatment Reverses Prematurity-Induced Disruption in Cortical Interneuron Population.

Development of cortical interneurons continues until the end of human pregnancy. Premature birth deprives the newborns from the supply of maternal estrogen and a secure intrauterine environment. Indeed, preterm infants suffer from neurobehavioral disorders. This can result from both preterm birth and associated postnatal complications, which might disrupt recruitment and maturation of cortical interneurons. We hypothesized that interneuron subtypes, including parvalbumin-positive (PV+), somatostatin-positive (SST+), calretinin-positive (CalR+), and neuropeptide Y-positive (NPY+) interneurons, were recruited in the upper and lower cortical layers in a distinct manner with advancing gestational age. In addition, preterm birth would disrupt the heterogeneity of cortical interneurons, which might be reversed by estrogen treatment. These hypotheses were tested by analyzing autopsy samples from premature infants and evaluating the effect of estrogen supplementation in prematurely delivered rabbits. The PV+ and CalR+ neurons were abundant, whereas SST+ and NPY+ neurons were few in cortical layers of preterm human infants. Premature birth of infants reduced the density of PV+ or GAD67+ neurons and increased SST+ interneurons in the upper cortical layers. Importantly, 17 ß-estradiol treatment in preterm rabbits increased the number of PV+ neurons in the upper cortical layers relative to controls at postnatal day 14 (P14) and P21 and transiently reduced SST population at P14. Moreover, protein and mRNA levels of Arx, a key regulator of cortical interneuron maturation and migration, were higher in estrogen-treated rabbits relative to controls. Therefore, deficits in PV+ and excess of SST+ neurons in premature newborns are ameliorated by estrogen replacement, which can be attributed to elevated Arx levels. Estrogen replacement might enhance neurodevelopmental outcomes in extremely preterm infants.SIGNIFICANCE STATEMENT Premature birth often leads to neurodevelopmental delays and behavioral disorders, which may be ascribed to disturbances in the development and maturation of cortical interneurons. Here, we show that preterm birth in humans is associated with reduced population of parvalbumin-positive (PV+) neurons and an excess of somatostatin-expressing interneurons in the cerebral cortex. More importantly, 17 ß-estradiol treatment increased the number of PV+ neurons in preterm-born rabbits, which appears to be mediated by an elevation in the expression of Arx transcription factor. Hence the present study highlights prematurity-induced reduction in PV+ neurons in human infants and reversal in their population by estrogen replacement in preterm rabbits. Because preterm birth drops plasma estrogen level 100-fold, estrogen replacement in extremely preterm infants might improve their developmental outcome and minimize neurobehavioral disorders.

Decreased response of interneurons in the medial prefrontal cortex to 5-HT1A receptor activation in the rat 6-hydroxydopamine Parkinson model.

This study examined the response of interneurons in the medial prefrontal cortex (mPFC) to 5-HT1A receptor agonist 8-OH-DPAT and change in expression of 5-HT1A receptor on glutamate decarboxylase 67 (GAD67)-positive neurons in rats with 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra pars compacta (SNc). Systemic administration of 5-HT1A receptor agonist 8-OH-DPAT dose-dependently inhibited the firing rate of the interneurons at all doses tested in sham-operated rats. In 6-OHDA-lesioned rats, 8-OH-DPAT, at the same doses, also inhibited the firing rate of the interneurons, whereas the inhibition was significant only at a high cumulative dose. Furthermore, injection of 8-OH-DPAT into the mPFC inhibited the interneurons in sham-operated rats, while having no effect on firing rate of the interneurons in 6-OHDA-lesioned rats. In contrast to sham-operated rats, SNc lesion reduced the expression of 5-HT1A receptor on GAD67-positive neurons in the prelimbic cortex, a sub-region of the mPFC. Our results indicate that degeneration of the nigrostriatal pathway leads to decreased response of mPFC interneurons to 5-HT1A receptor activation, which attributes to the down-regulation of 5-HT1A receptor expression in these interneurons.

Effects of chronic nicotine on the autocrine regulation of pancreatic cancer cells and pancreatic duct epithelial cells by stimulatory and inhibitory neurotransmitters.

Pancreatic ductal adenocarcinoma (PDAC) has a mortality rate near 100%. Smoking is a documented risk factor. However, the mechanisms of smoking-associated pancreatic carcinogenesis are poorly understood. We have shown that binding of nicotine to nicotinic acetylcholine receptors (nAChRs) expressing subunits α7, α3 and α5 in PDAC and pancreatic duct epithelial cells in vitro triggered the production of the neurotransmitters noradrenaline and adrenaline by these cells. In turn, this autocrine catecholamine loop significantly stimulated cell proliferation via cyclic adenosine 3',5'-monophosphate-dependent signaling downstream of beta-adrenergic receptors. However, the observed responses only represent acute cellular reactions to single doses of nicotine whereas nicotine exposure in smokers is chronic. Using the PDAC cell lines BxPC-3 and Panc-1 and immortalized pancreatic duct epithelial cell line HPDE6-C7, our current experiments reveal a significant sensitization of the nAChR-driven autocrine catecholamine regulatory loop in cells pre-exposed to nicotine for 7 days. The resulting increase in catecholamine production was associated with significant inductions in the phosphorylation of signaling proteins ERK, CREB, Src and AKT, upregulated protein expression of nAChR subunits α3, α4, α5 and α7 and increased responsiveness to nicotine in 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide and cell migration assays. All three cell lines produced the inhibitory neurotransmitter γ-aminobutyric acid, an activity inhibited by gene knockdown of the α4ß2nAChR and suppressed by chronic nicotine via receptor desensitization. All of the observed adverse effects of chronic nicotine were reversed by treatment of the cells with γ-aminobutyric acid, suggesting the potential usefulness of this agent for the improvement of PDAC intervention strategies in smokers.

Novel object recognition ability in female mice following exposure to nanoparticle-rich diesel exhaust.

Recently, our laboratory reported that exposure to nanoparticle-rich diesel exhaust (NRDE) for 3 months impaired hippocampus-dependent spatial learning ability and up-regulated the expressions of memory function-related genes in the hippocampus of female mice. However, whether NRDE affects the hippocampus-dependent non-spatial learning ability and the mechanism of NRDE-induced neurotoxicity was unknown. Female BALB/c mice were exposed to clean air, middle-dose NRDE (M-NRDE, 47 µg/m(3)), high-dose NRDE (H-NRDE, 129 µg/m(3)), or filtered H-NRDE (F-DE) for 3 months. We then investigated the effect of NRDE exposure on non-spatial learning ability and the expression of genes related to glutamate neurotransmission using a novel object recognition test and a real-time RT-PCR analysis, respectively. We also examined microglia marker Iba1 immunoreactivity in the hippocampus using immunohistochemical analyses. Mice exposed to H-NRDE or F-DE could not discriminate between familiar and novel objects. The control and M-NRDE-exposed groups showed a significantly increased discrimination index, compared to the H-NRDE-exposed group. Although no significant changes in the expression levels of the NMDA receptor subunits were observed, the expression of glutamate transporter EAAT4 was decreased and that of glutamic acid decarboxylase GAD65 was increased in the hippocampus of H-NRDE-exposed mice, compared with the expression levels in control mice. We also found that microglia activation was prominent in the hippocampal area of the H-NRDE-exposed mice, compared with the other groups. These results indicated that exposure to NRDE for 3 months impaired the novel object recognition ability. The present study suggests that genes related to glutamate metabolism may be involved in the NRDE-induced neurotoxicity observed in the present mouse model.

[Changes in expression levels of PV, GAD67 and KCC2 in the brain tissue of rats with schizophrenia induced by MK-801].

OBJECTIVE: To study changes in the expression levels of parvalbumin (PV), glutamate decarboxylase 67 (GAD67) and K+-Cl- cotransporter 2 (KCC2) in the brain tissue of rats with schizophrenia (SZ) induced by dizocilpine (MK-801), and to investigate the mechanism involving gamma-aminobutyric acid (GABA) by which NMDA receptor blocker induces SZ in the perinatal period. METHODS: Thirty-six neonatal male Sprague-Dawley rats were randomly assigned to two batches on postnatal day 6. Each batch was divided into normal control (treated by 0.9% normal saline), SZ-development model (treated by subcutaneous injection of 0.1 mg/kg MK-801 on postnatal days 7-10; bid), and SZ-chronic medication model groups (treated by intraperitoneal injection of 0.2 mg/kg MK-801 on postnatal days 47-60; qd). On postnatal day 63, the brain tissue of the first batch of rats was obtained and then fixed with paraform for histological sections; expression levels of PV and GAD67 in the medial prefrontal cortex (mPFC) and hippocampus CA1 were measured by immunohistochemistry. Simultaneously, the second batch of rats was sacrificed and the mPFC and hippocampus were obtained and homogenized; expression levels of KCC2 in the mPFC and hippocampus were measured by Western blot. RESULTS: Expression levels of PV and GAD67 in the mPFC and hippocampus CA1 were significantly lower in the SZ-development and chronic medication model groups than in the normal control group (P<0.05). Expression levels of KCC2 in the mPFC and hippocampus were significantly lower in the SZ-development model group than in the SZ-chronic medication model and normal control groups (P<0.05). CONCLUSIONS: The expression changes of PV and GAD67 in SZ can be simulated using the SZ development model induced by MK-801, which might affect the development of the GABA system in the PFC and hippocampus by downregulating KCC2 expression.

Kalirin-7, an important component of excitatory synapses, is regulated by estradiol in hippocampal neurons.

Estradiol enhances the formation of dendritic spines and excitatory synapses in hippocampal neurons in vitro and in vivo, but the underlying mechanisms are not fully understood. Kalirin-7 (Kal7), the major isoform of Kalirin in the adult hippocampus, is a Rho GDP/GTP exchange factor localized to postsynaptic densities. In the hippocampus, both Kal7 and estrogen receptor α (ERα) are highly expressed in a subset of interneurons. Over-expression of Kal7 caused an increase in spine density and size in hippocampal neurons. To determine whether Kalirin might play a role in the effects of estradiol on spine formation, Kal7 expression was examined in the hippocampus of ovariectomized rats. Estradiol replacement increased Kal7 staining in both CA1 pyramidal neurons and interneurons in ovariectomized rats. Estradiol treatment of cultured hippocampal neurons increased Kal7 levels at the postsynaptic side of excitatory synapses and increased the number of excitatory synapses along the dendrites of pyramidal neurons. These increases were mediated via ERα because a selective ERα agonist, but not a selective ERß agonist, caused a similar increase in both Kal7 levels and excitatory synapse number in cultured hippocampal neurons. When Kal7 expression was reduced using a Kal7-specific shRNA, the density of excitatory synapses was reduced and estradiol was no longer able to increase synapse formation. Expression of exogenous Kal7 in hippocampal interneurons resulted in decreased levels of GAD65 staining. Inhibition of GABAergic transmission with bicuculline produced a robust increase in Kal7 expression. These studies suggest Kal7 plays a key role in the mechanisms of estradiol-mediated synaptic plasticity.

High titre of antiglutamic acid decarboxylase autoantibody is a strong predictor of the development of thyroid autoimmunity in patients with type 1 diabetes and latent autoimmune diabetes in adults.

OBJECTIVE: Type 1 diabetes mellitus (T1DM) is frequently associated with autoimmune thyroid diseases (AITD), but little is known about the risk of AITD in latent autoimmune diabetes in adults (LADA). We evaluated the genetic and immunological factors involved in the development of thyroid autoimmunity in patients with LADA and T1DM. PATIENTS AND MEASUREMENTS: One hundred and ninety T1DM and 135 LADA patients were recruited in the study. Thyroid peroxidase antibody (TPOAb), thyroglobulin antibody (TGAb), glutamic acid decarboxylase antibody (GADA) and thyroid function were measured. The cytotoxic-lymphocyte-associated antigen-4 (CTLA-4) +49A/G and CT60 polymorphisms and the human leucocyte antigen (HLA)-DQA1-DQB1 genotype were determined. RESULTS: The prevalence of thyroid antibodies (TGAb and/or TPOAb) and thyroid dysfunction was 27·4% and 9·5% in patients with T1DM, and 21·5% and 11·1% in patients with LADA. Thyroid-antibody-positive T1DM patients had higher frequencies of GADA and HLA-DQA1*03-DQB1*0401 haplotypes than thyroid-antibody negatives (P < 0·05). Thyroid-antibody-positive LADA patients had higher GADA titre, lower C-peptide levels and higher frequencies of HLA-DQA1*03-DQB1*0401 haplotypes (P < 0·05). The CTLA-4 +49A/G and CT60 polymorphism was associated with T1DM complicated with thyroid autoimmunity (OR = 2·33 and 2·54). Logistic regression revealed that only high-titre GADA was associated with development of thyroid autoimmunity in patients with T1DM and LADA (OR = 3·50 and 3·10, respectively), and the presence of thyroid antibody predicted high risk for thyroid dysfunction in patients with T1DM and LADA (OR = 9·25 and 10·70, respectively). CONCLUSION: Regular screening of thyroid antibody and function are recommended, especially in patients with T1DM and LADA with high GADA titre.

[Discovering a new functional neurogenic zone: the vestibular nuclei of the brainstem]. / Une nouvelle zone de neurogenèse fonctionnelle: les noyaux vestibulaires du tronc cérébral.

The adult mammal brain is mostly considered as non-neurogenic, except in the subventricular zone of the lateral ventricles and the subgranular zone of the dentate gyrus, where ongoing neurogenesis occurs. However, anti-neurogenic influences can be removed in pathological conditions or after specific injury. That is what happens in a model of unilateral vestibular neurectomy (UVN) that mimics human pathology in adult cats. We showed for the first time that a UVN promoted an intense reactive cell proliferation in the deafferented vestibular nuclei located in the brainstem. The new cells survived up to one month, differentiated into glial cells - microglia or astrocytes - or GABAergic neurons, so highlighting a GABAergic neurogenesis. Surprisingly, we further showed that post-UVN reactive cell proliferation contributed successfully to fine restoration of vestibular posturo-locomotor functions. In conclusion, these pioneering studies bring new pieces of a promising puzzle in both stem cell and vestibular therapy domains.

The relationship between dlx and gad1 expression indicates highly conserved genetic pathways in the zebrafish forebrain.

The Dlx genes encode a family of transcription factors important for the development of the vertebrate forebrain. These genes have very similar expression domains during the development of the telencephalon in mice and play a role in gamma-aminobutyric acid (GABAergic) interneuron differentiation. We have used triple fluorescent in situ hybridization to study the relative expression domains of the dlx and gad1 genes in the zebrafish telencephalon and diencephalon. We also generated transgenic zebrafish with regulatory elements from the zebrafish dlx1a/2a locus. The zebrafish dlx regulatory elements recapitulated dlx expression in the forebrain and mimicked the relationship between the expression of the dlx genes and gad1. Finally, we show that a putative enhancer located downstream of dlx2b can also activate reporter gene expression in a tissue-specific manner similar to endogenous dlx2b expression. Our results indicate the dlx genes are regulated by an evolutionarily conserved genetic pathway and may play a role in GABAergic interneuron differentiation in the zebrafish forebrain.

Differential HLA Association of GAD65 and IA2 Autoantibodies in North Indian Type 1 Diabetes Patients.

The human leucocyte antigen (HLA) association with type 1 diabetes (T1D) is well known but there are limited studies investigating the association between ß-cell autoantibodies and HLA genes. We evaluated the prevalence of GAD65 and IA-2 autoantibodies (GADA and IA2A) in 252 T1D patients from North India and investigated the genetic association of GADA and IA2A with HLA class I and class II genes/haplotypes. GADA and IA2A were detected in 50.79% and 15.87% of T1D patients, respectively, while only 8.73% had both GADA and IA2A. HLA-DRB1∗03 was observed to be significantly higher in GADA+ T1D patients as compared to GADA- (91.41% vs. 66.13%, Bonferroni-corrected P (P c) = 1.11 × 10-5; OR = 5.45; 95% CI: 2.67-11.08). Similarly, HLA-DQB1∗02 was found to be significantly increased in GADA+ patients (94.53%, P c = 2.19 × 10-5; OR = 6.27; 95% CI: 2.7-14.49) as compared to GADA- (73.39%). The frequencies of HLA-DRB1∗04 and DQB1∗03 were increased in IA2A+ patients (45.0% and 52.5%, respectively) as compared to that in IA2A- (25.94% and 33.96%, respectively). Further, the frequency of DRB1∗03-DQB1∗02 haplotype was found to be significantly increased in GADA+ T1D patients as compared to GADA- (60.55% vs. 41.94%, P = 3.94 × 10-5; OR = 2.13; 95%CI = 1.49-3.03). Similarly, HLA-DRB1∗04-DQB1∗03 haplotype was found to be significantly increased in IA2A+ T1D patients compared to IA2A- patients (22.5% vs. 12.97%; P = 0.041; OR = 1.95; 95%CI = 1.08-3.52). None of the HLA class I genes (HLA-A, B, and Cw) was found to be associated with GADA or IA2A in people with T1D. Our findings suggest that HLA-DRB1∗03/DQB1∗02 and HLA-DRB1∗04/DQB1∗03 might play an important role in the development of GADA and IA2A, respectively.

Two classes of GABAergic neurons in the inferior colliculus.

The inferior colliculus (IC) is unique, having both glutamatergic and GABAergic projections ascending to the thalamus. Although subpopulations of GABAergic neurons in the IC have been proposed, criteria to distinguish them have been elusive and specific types have not been associated with specific neural circuits. Recently, the largest IC neurons were found to be recipients of somatic terminals containing vesicular glutamate transporter 2 (VGLUT2). Here, we show with electron microscopy that VGLUT2-positive (VGLUT2(+)) axonal terminals make axosomatic synapses on IC neurons. These terminals contain only VGLUT2 even though others in the IC have VGLUT1 or both VGLUT1 and 2. We demonstrate that there are two types of GABAergic neurons: larger neurons with VGLUT2(+) axosomatic endings and smaller neurons without such endings. Both types are present in all subdivisions of the IC, but larger GABAergic neurons with VGLUT2(+) axosomatic terminals are most prevalent in the central nucleus. The GABAergic tectothalamic neurons consist almost entirely of the larger cells surrounded by VGLUT2(+) axosomatic endings. Thus, two types of GABAergic neurons in the IC are defined by different synaptic organization and neuronal connections. Larger tectothalamic GABAergic neurons are covered with glutamatergic axosomatic synapses that could allow them to fire rapidly and overcome a slow membrane time constant; their axons may be the largest in the brachium of the IC. Thus, large GABAergic neurons could deliver IPSPs to the medial geniculate body before EPSPs from glutamatergic IC neurons firing simultaneously.

New GABAergic interneurons in the adult neocortex and striatum are generated from different precursors.

Ongoing neurogenesis in the adult mammalian dentate gyrus and olfactory bulb is generally accepted, but its existence in other adult brain regions is highly controversial. We labeled newly born cells in adult rats with the S-phase marker bromodeoxyuridine (BrdU) and used neuronal markers to characterize new cells at different time points after cell division. In the neocortex and striatum, we found BrdU-labeled cells that expressed each of the eight neuronal markers. Their size as well as staining for gamma-aminobutyric acid (GABA), glutamic acid decarboxylase 67, calretinin and/or calbindin, suggest that new neurons in both regions are GABAergic interneurons. BrdU and doublecortin-immunoreactive (BrdU+/DCX+) cells were seen within the striatum, suggesting migration of immature neurons from the subventricular zone. Surprisingly, no DCX+ cells were found within the neocortex. NG2 immunoreactivity in some new neocortical neurons suggested that they may instead be generated from the NG2+ precursors that reside within the cortex itself.

Presence of human herpes virus 6 DNA exclusively in temporal lobe epilepsy brain tissue of patients with history of encephalitis.

Temporal lobe epilepsy (TLE) is frequently associated with mesial temporal sclerosis (MTS). Many etiologic aspects of TLE are still unresolved. Here, we aimed to analyze the presence of human herpes virus 6 (HHV-6) DNA in distinct TLE pathologies. Nested polymerase chain reaction (PCR) in surgical tissue from 38 pharmaco-resistant TLE patients and 10 autopsy controls revealed HHV-6 DNA in 55.6% of the TLE patients with a history of encephalitis, involving MTS and gliotic hippocampi without substantial neurodegeneration, but not in lesion-associated TLE or nonlesional MTS with or without a history of complex febrile seizures (CFS). HHV-6 protein was present in only one patient's tissue. Our data argue against HHV-6 as a major local pathogenetic factor in MTS hippocampi after CFS. The high detection rate of HHV-6 DNA suggests a potential pathogenetic role of HHV-6 in TLE patients with a history of encephalitis.

Cell-Type-Specific Whole-Brain Direct Inputs to the Anterior and Posterior Piriform Cortex.

The piriform cortex (PC) is a key brain area involved in both processing and coding of olfactory information. It is implicated in various brain disorders, such as epilepsy, Alzheimer's disease, and autism. The PC consists of the anterior (APC) and posterior (PPC) parts, which are different anatomically and functionally. However, the direct input networks to specific neuronal populations within the APC and PPC remain poorly understood. Here, we mapped the whole-brain direct inputs to the two major neuronal populations, the excitatory glutamatergic principal neurons and inhibitory γ-aminobutyric acid (GABA)-ergic interneurons within the APC and PPC using the rabies virus (RV)-mediated retrograde trans-synaptic tracing system. We found that for both types of neurons, APC and PPC share some similarities in input networks, with dominant inputs originating from the olfactory region (OLF), followed by the cortical subplate (CTXsp), isocortex, cerebral nuclei (CNU), hippocampal formation (HPF) and interbrain (IB), whereas the midbrain (MB) and hindbrain (HB) were rarely labeled. However, APC and PPC also show distinct features in their input distribution patterns. For both types of neurons, the input proportion from the OLF to the APC was higher than that to the PPC; while the PPC received higher proportions of inputs from the HPF and CNU than the APC did. Overall, our results revealed the direct input networks of both excitatory and inhibitory neuronal populations of different PC subareas, providing a structural basis to analyze the diverse PC functions.

Effect of Immune Activation during Early Gestation or Late Gestation on Inhibitory Markers in Adult Male Rats.

People with schizophrenia exhibit deficits in inhibitory neurons and cognition. The timing of maternal immune activation (MIA) may present distinct schizophrenia-like phenotypes in progeny. We investigated whether early gestation [gestational day (GD) 10] or late gestation (GD19) MIA, via viral mimetic polyI:C, produces deficits in inhibitory neuron indices (GAD1, PVALB, SST, SSTR2 mRNAs) within cortical, striatal, and hippocampal subregions of male adult rat offspring. In situ hybridisation revealed that polyI:C offspring had: (1) SST mRNA reductions in the cingulate cortex and nucleus accumbens shell, regardless of MIA timing; (2) SSTR2 mRNA reductions in the cortex and striatum of GD19, but not GD10, MIA; (3) no alterations in cortical or striatal GAD1 mRNA of polyI:C offspring, but an expected reduction of PVALB mRNA in the infralimbic cortex, and; (4) no alterations in inhibitory markers in hippocampus. Maternal IL-6 response negatively correlated with adult offspring SST mRNA in cortex and striatum, but not hippocampus. These results show lasting inhibitory-related deficits in cortex and striatum in adult offspring from MIA. SST downregulation in specific cortical and striatal subregions, with additional deficits in somatostatin-related signalling through SSTR2, may contribute to some of the adult behavioural changes resulting from MIA and its timing.

Pedunculopontine and laterodorsal tegmental nuclei contain distinct populations of cholinergic, glutamatergic and GABAergic neurons in the rat.

The pedunculopontine tegmental nucleus (PPTg) and laterodorsal tegmental nucleus (LDTg) provide cholinergic afferents to several brain areas. This cholinergic complex has been suggested to play a role in sleep, waking, motor function, learning and reward. To have a better understanding of the neurochemical organization of the PPTg/LDTg we characterized the phenotype of PPTg/LDTg neurons by determining in these cells the expression of transcripts encoding choline acetyltransferase (ChAT), glutamic acid decarboxylase (GAD) or the vesicular glutamate transporters (vGluT1, vGluT2 and vGluT3). Within the PPTg/LDTg complex we found neurons expressing ChAT, vGluT2 or GAD transcripts, these neuronal phenotypes were intermingled, but not homogeneously distributed within the PPTg or LDTg. Previous studies suggested the presence of either glutamate or gamma-aminobutyric acid (GABA) immunolabeling in a large number of PPTg/LDTg cholinergic neurons, leading to the widespread notion that PPTg/LDTg cholinergic neurons co-release acetylcholine together with either glutamate or GABA. To assess the glutamatergic or GABAergic nature of the PPTg/LDTg cholinergic neurons, we combined in situ hybridization (to detect vGluT2 or GAD transcripts) and immunohistochemistry (to detect ChAT), and found that over 95% of all PPTg/LDTg cholinergic neurons lack transcripts encoding either vGluT2 mRNA or GAD mRNA. As the vast majority of PPTg/LDTg cholinergic neurons lack transcripts encoding essential proteins for the vesicular transport of glutamate or for the synthesis of GABA, co-release of acetylcholine with either glutamate or GABA is unlikely to be a major factor in the interactions between acetylcholine, glutamate and GABA at the postsynaptic site.