Somatostatin mRNA Research Articles

Olfactory placode generates a diverse population of neurons expressing GnRH, somatostatin mRNA, neuropeptide Y, or calbindin in the chick forebrain.

The olfactory placode (OP) of vertebrates generates several classes of migrating cells, including hypothalamic gonadotropin-releasing hormone (GnRH)-producing neurons, which play essential roles in the reproduction system. Previous studies using OP cell labeling have demonstrated that OP-derived non-GnRH cells enter the developing forebrain; however, their final fates and phenotypes are less well understood. In chick embryos, a subpopulation of migratory cells from the OP that is distinct from GnRH neurons transiently expresses somatostatin (SS). We postulated that these cells are destined to develop into brain neurons. In this study, we examined the expression pattern of SS mRNA in the olfactory-forebrain region during development, as well as the destination of OP-derived migratory cells, including SS mRNA-expressing cells. Utilizing the Tol2 genomic integration system to induce long-term fluorescent protein expression in OP cells, we found that OP-derived migratory cells labeled at embryonic day (E) 3 resided in the olfactory nerve and medial forebrain at E17-19. A subpopulation of green fluorescent protein (GFP)-labeled GnRH neurons that remained in the olfactory nerve was considered to comprise terminal nerve neurons. In the forebrain, GFP-labeled cells showed a distribution pattern similar to that of GnRH neurons. A large proportion of GFP-labeled cells expressed the mature neuronal marker NeuN. Among the GFP-labeled cells, the percentage of GnRH neurons was low, while the remaining GnRH-negative neurons either expressed SS mRNA, neuropeptide Y, or calbindin D-28k or did not express any of them. These results indicate that a diverse population of OP-derived neuronal cells, other than GnRH neurons, integrates into the chick medial forebrain.

The Distinct Characteristics of Somatostatin Neurons in the Human Brain.

Somatostatin cells are frequently described as a major population of GABAergic neurons in the cerebral cortex. In this study, we performed a comprehensive analysis of their molecular expression, morphological features, and laminar distribution. We provided a detailed description of somatostatin neurons in the human prefrontal cortex, including their proportion in the total neuron population, laminar distribution, neurotransmitter phenotype, as well as their molecular and morphological characteristics using immunofluorescence and RNAscope in situ hybridization. We found that somatostatin neurons comprise around 7% of neocortical neurons in the human Brodmann areas 9 and 14r, without significant difference between the two regions. Somatostatin cells were NeuN positive and synthesized vesicular GABA transporter and glutamate decarboxylase 1 and 2, confirming their neuronal nature and GABAergic phenotype. Somatostatin cells in the upper cortical layers were small, had a high expression of somatostatin mRNA, a relatively low expression of somatostatin peptide, and co-expressed calbindin. In the lower cortical layers, somatostatin cells were larger with complex somato-dendritic morphology, typically showed a lower expression of somatostatin mRNA and a high expression of somatostatin peptide, and co-expressed neuronal nitric oxide synthase (nNOS) and neuropeptide Y (NPY), but not calbindin. Somatostatin neurons in the white matter co-expressed MAP2. Based on their somato-dendritic morphology, cortical somatostatin neurons could be classified into at least five subtypes. The somatostatin neurons of the human prefrontal cortex show remarkable morphological and molecular complexity, which implies that they have equally complex and distinct functions in the human brain.

Density of Somatostatin Neurons in the Dorsolateral Prefrontal Cortex of Schizophrenia Subjects

Cognitive dysfunction in schizophrenia is associated with altered GABA signaling markers in the prefrontal cortex (PFC). For example, levels of somatostatin (SST) mRNA, expressed in a subset of GABA neurons, are markedly lower in PFC gray matter in schizophrenia. The density of SST mRNA+ neurons is also lower in schizophrenia. However, whether schizophrenia is associated with fewer SST neurons or a failure of certain SST neurons to express detectable levels of SST mRNA remains unknown.

Leptin Modulates the Response of Brown Adipose Tissue to Negative Energy Balance: Implication of the GH/IGF-I Axis.

The growth hormone (GH)/insulin-like growth factor I (IGF-I) axis is involved in metabolic control. Malnutrition reduces IGF-I and modifies the thermogenic capacity of brown adipose tissue (BAT). Leptin has effects on the GH/IGF-I axis and the function of BAT, but its interaction with IGF-I and the mechanisms involved in the regulation of thermogenesis remains unknown. We studied the GH/IGF-I axis and activation of IGF-I-related signaling and metabolism related to BAT thermogenesis in chronic central leptin infused (L), pair-fed (PF), and control rats. Hypothalamic somatostatin mRNA levels were increased in PF and decreased in L, while pituitary GH mRNA was reduced in PF. Serum GH and IGF-I concentrations were decreased only in PF. In BAT, the association between suppressor of cytokine signaling 3 and the IGF-I receptor was reduced, and phosphorylation of the IGF-I receptor increased in the L group. Phosphorylation of Akt and cyclic AMP response element binding protein and glucose transporter 4 mRNA levels were increased in L and mRNA levels of uncoupling protein-1 (UCP-1) and enzymes involved in lipid anabolism reduced in PF. These results suggest that modifications in UCP-1 in BAT and changes in the GH/IGF-I axis induced by negative energy balance are dependent upon leptin levels.

Somatostatin receptor subtype 4 protects against indomethacin-induced gastrointestinal mucosal injury in mouse

Abstract Despite recent advancements in the pharmacologic and endoscopic management of gastrointestinal ulceration, it is still associated with high mortality. Therefore, it is important to understand mucosal protective mechanisms. Somatostatin, present in the endocrine cells & sensory nerves, and its analogues reduce mucosal blood flow, pepsin and gastric acid secretion. Its receptor subtype 4 (sst4) mediates anti-inflammatory & analgesic actions, as discovered by our group. Therefore, our aim was to investigate its role in gastrointestinal injury. We treated sst4 gene-deleted (sst4−/−) and wildtype mice with indomethacine (IDM; 35 mg/kg) or vehicle s.c.. Gastric & small intestine lesions were evaluated after 4 & 48 h macroscopically & microscopically. Plasma protein extravasation & myeloperoxidase (MPO) activity were assessed by ex vivo imaging, TNFα, IL-1, COX2, sst4 & somatostatin mRNA levels by qPCR. Gastric (4h) and small intestinal macroscopic (24 h) lesions, MPO activity (24, 48 h), and plasma extravasation (30 min) were significantly greater in IDM-treated sst4−/− mice, while histopathological scores & small intestinal length did not show strain difference. TNFα, IL-1, COX2 significantly upregulated after 4h in the stomach of sst4−/− mice, and the same pattern was observable in wildtype mice at later timepoint (48 h). These results provide evidence for the protective role of the sst4 receptor against IDM-induced gastrointestinal mucosal injury. This suggests the safety of sst4 agonist drug candidates in the gastrointestinal tract, and even their potential use for gastroprotective indication. Supported by: GINOP-2.3.2-15-2016-00050 – PEPSYS, GINOP-2.3.2 STAY ALIVE, EFOP-3.6.2-16-2017-00008, ÚNKP-19-3-III-PTE-211

Expression and Protective Functions of the sst4 Somatostatin Receptor in the Indomethacin‐Induced Gastrointestinal Mucosal Injury Model of the Mouse

IntroductionGastrointestinal ulceration is associated with high mortality despite advanced management, therefore, understanding of the mucosal protective mechanisms is important. Somatostatin is present in the endocrine cells and sensory nerves, and its analogues were shown to reduce mucosal blood flow, pepsin and gastric acid secretion. Our group discovered that its anti‐inflammatory and analgesic actions are exerted via the somatostatin receptor subtype 4 (sst4). Therefore, we investigated its expression and role in gastrointestinal injury.MethodsSst4 gene‐deleted (sst4−/−) and wildtype mice were treated s.c. with indomethacine (IDM; 35 mg/kg) or vehicle. RNA scope in situ hybridization was performed to localize, qPCR to quantify sst4 and somatostatin mRNA. Macroscopic and microscopic lesions in the stomach and small intestine were evaluated after 4 and 48 h. Plasma protein extravasation was measured by fluorescent, neutrophil myeloperoxidase (MPO) activity bioluminescent ex vivo imaging, TNFα, IL‐1 and COX2 mRNA by qPCR.ResultsSomatostatin mRNA is present in the neuroendocrine D‐cells and significantly upregulated 48 h after IDM. Sst4 mRNA in located in the myenteric plexus neurons, it’s expression did not change in response to mucosal injury.IDM‐induced gastric and small intestinal macroscopic lesions (4, 24 h), gastric MPO activity (24, 48 h) and plasma extravasation (30 min) were significantly greater in sst4−/− mice. TNFα, IL‐1, COX2 expressions significantly increased after 4 h in the stomach of sst4−/− mice, but in wild‐types the same pattern was observed only later, at the 48 h time‐point. IDM‐evoked histopathological alterations and small intestinal length reduction were not altered by the sst4 deletion.ConclusionThis is the first evidence for sst4 expression in myenteric plexus neurons and its protective roles against chemically‐induced gastrointestinal mucosal damage and inflammation. This suggests the safety of sst4 agonist drug candidates in the gastrointestinal tract, and even their potential use for gastroprotective indication.Support or Funding InformationSupported by: GINOP‐2.3.2‐15‐2016‐00050 – PEPSYS, GINOP‐2.3.2 STAY ALIVE, EFOP‐3.6.2‐16‐2017‐00008, ÚNKP‐19‐3‐III‐PTE‐211

Effects of coated cysteamine on growth performance, carcass characteristics, meat quality and lipid metabolism in finishing pigs

This study was conducted to evaluate the effects of coated cysteamine (CCS) on growth performance, carcass characteristics, meat quality, and lipid metabolism in finishing pigs. A total of 160 pigs (62.12 ± 1.43 kg body weight) were randomly divided into 4 groups, with 4 pens per group and 10 pigs (5 gilts and 5 barrows) per pen. Pigs were fed a basal diet supplemented with 0, 100, 200, or 400 mg/kg CCS, corresponding to 0, 27, 54, or 108 mg/kg cysteamine, respectively. All pigs were given free access to feed and water for forty days. Average daily gain, average daily feed intake and feed to gain ratio were not affected by the CCS treatment. Supplemental CCS increased carcass lean ratio linearly (P < 0.05) and longissmus muscle area quadratically (P < 0.05), and the water-holding capacity as measured by 24 h drip loss was quadratically affected (P < 0.05) by CCS supplementation. The activity of hormone-sensitive lipase in subcutaneous adipose tissue was linearly increased (P < 0.001), while fatty acid synthase activity was linearly decreased (P < 0.05). Pigs fed CCS had a lower serum triacylglycerol (P < 0.001) level and lipase activity (P < 0.001), lower total cholesterol (P < 0.05) level, and a higher total protein (P < 0.001) and free fatty acids (P < 0.01) levels. In addition, CCS quadratically increased serum growth hormone (GH) (P < 0.001) level and quadratically affected somatostatin (SS) (P < 0.05) level. Consistently, significant effects of dietary CCS were observed on the mRNA expression of GH and SS in the anterior pituitary gland and the hypothalamus, respectively, where the intermediate CCS level showed the highest GH mRNA level and lowest SS mRNA level. The results indicate that dietary supplementation of CCS has beneficial effects on carcass characteristics and meat quality, and affects lipid metabolism in finishing pigs.

Regulation of somatostatin expression by vitamin D3 and valproic acid in human adipose-derived mesenchymal stem cells

BackgroundAdipose-derived mesenchymal stem cells (ADMSC) are non-haematopoietic, fibroblast-like multipotent progenitor cells. They have the potential for trilineage (adipocyte, chondrocyte and osteocyte) differentiation as well as differentiation into endocrine pancreatic progenitors. In diabetic or cancer therapy, somatostatin (SST) expression plays a vital role. Small molecules such as valproic acid (VPA) and micronutrients like vitamin D3 have differentiation potential in ADMSC. Therefore, the aim of this study was to investigate the role of vitamin D3 machinery and its metabolic enzymes in ADMSC. Furthermore, the reprogramming effect of vitamin D3 and VPA was evaluated on somatostatin expression in pancreatic lineage differentiation.MethodsADMSC were characterised based on their cell surface marker profile using flow cytometry. Specific adipogenic and osteogenic differentiation protocols were used in this study. Gene expression of several pluripotent, endodermal, pancreatic progenitor and pancreatic endocrine lineage markers were investigated in native ADMSC and after stimulation with different concentration of vitamin D3 for five consecutive days (0, 50, 100, 150 nM) and VPA (0.5, 1, 1.5, 2 mM) by real-time PCR. Furthermore, somatostatin expression was confirmed with ELISA and immunocytochemistry.ResultsIn ADMSC, the expression of somatostatin mRNA, the vitamin D receptor (VDR) and its metabolising enzymes 1 α-Hydroxylase, 24-Hydroxylase and 25-Hydroxylase were detected. Upon stimulation with vitamin D3, nuclear translocation of vitamin D receptor (VDR) was observed. Interestingly, the presence of vitamin D3 reduced the transcription of the somatostatin gene. By contrast, VPA treatment of cultivated ADMSC showed enhancing effect on somatostatin gene expression. No other pluripotent, endodermal, pancreatic progenitor or pancreatic endocrine lineage mRNA expression was modulated under the influence of vitamin D3 and VPA.ConclusionHuman ADMSC carry the VDR. The vitamin D metabolising enzyme 25-Hydroxylase responded to the addition of vitamin D3. Moreover, our results demonstrate that somatostatin expression in ADMSC is constitutive, partially secreted and regulated by vitamin D3 and VPA.

Lack of Brain Insulin Receptor Substrate-1 (IRS1) Causes Growth Retardation via Growth Hormone Releasing Hormone

Background and Purpose: We previously revealed that systemic Irs1 deficient mice showed growth retardation, indicating that Irs1 plays an important role in growth. Irs1 is main substrate of not only insulin receptor (IR) but also IGF-1 receptor (IGF-1R) tyrosine kinase. Recently, brain specific IGF-1 receptor knockout mice were reported to become growth impairment. We hypothesized that brain Irs1 may regulate growth via IGF-1. Methods: We generated NIrs1KO mice and analyzed growth related parameter and immunohistochemistry (IHC) study. Result: The body weight of NIrs1KO mice was significantly smaller than control mice. NIrs1KO mice also showed shorter body length and bone length. On the other hand food intake and gene expression of appetite-related neuropeptides (POMC, AgRP and NPY) were not different between two groups. We next confirmed that plasma IGF-1 and growth hormone (GH) levels of NIrs1KO mice were lower than control mice. Hypothalamic GHRH mRNA expression levels of NIrs1KO mice were also lower than those of control mice although hypothalamic somatostatin mRNA expression was not different between two groups. These data strongly suggest that lack of brain Irs1 causes growth retardation via GHRH. IHC study indicated that Irs1 colocalized with GHRH neuron in hypothalamus. We next investigated the IGF-1-induced proliferation in hypothalamic neural cells by using siIrs1. Downregulation of Irs1 significantly inhibited IGF-1-induced neural cell proliferation. Conclusion: Our results suggest that IGF-1-Irs1 pathway promotes the proliferation of GHRH neurons and regulate growth. Disclosure T. Hayashi: None. N. Kubota: None. T. Kubota: None. T. Kadowaki: Consultant; Self; Novo Nordisk A/S, AstraZeneca, Merck Sharp &amp; Dohme Corp.. Research Support; Self; Kissei Pharmaceutical Co., Ltd., Taisho Pharmaceutical Co., Ltd., Sanofi, Kyowa Hakko Kirin Co., Ltd., Novo Nordisk A/S, Astellas Pharma, Daiichi Sankyo Company, Limited, Takeda, Mitsubishi Tanabe Pharma Corporation, Ono Pharmaceutical Co., Ltd., Merck Sharp &amp; Dohme Corp., Nippon Boehringer Ingelheim Co. Ltd.. Speaker's Bureau; Self; Astellas Pharma, AstraZeneca, Merck Sharp &amp; Dohme Corp., Ono Pharmaceutical Co., Ltd., Takeda, Eli Lilly and Company, Nippon Boehringer Ingelheim Co. Ltd., Novo Nordisk A/S.

A PCR-Based Method for RNA Probes and Applications in Neuroscience.

In situ hybridization (ISH) is a powerful technique that is used to detect the localization of specific nucleic acid sequences for understanding the organization, regulation, and function of genes. However, in most cases, RNA probes are obtained by in vitro transcription from plasmids containing specific promoter elements and mRNA-specific cDNA. Probes originating from plasmid vectors are time-consuming and not suitable for the rapid gene mapping. Here, we introduce a simplified method to prepare digoxigenin (DIG)-labeled non-radioactive RNA probes based on polymerase chain reaction (PCR) amplification and applications in free-floating mouse brain sections. Employing a transgenic reporter line, we investigate the expression of the somatostatin (SST) mRNA in the adult mouse brain. The method can be applied to identify the colocalization of SST mRNA and proteins including corticotrophin-releasing hormone (CRH) and protein kinase C delta type (PKC-δ) using double immunofluorescence, which is useful for understanding the organization of complex brain nuclei. Moreover, the method can also be incorporated with retrograde tracing to visualize the functional connection in the neural circuitry. Briefly, the PCR-based method for non-radioactive RNA probes is a useful tool that can be substantially utilized in neuroscience studies.

The influence of benazepril and amlodipine on the expression of secretin and somatostatin in spontaneously hypertensive rats

Investigate the influence of benazepril and amlodipine on the expression of secretin (PZ) and somatostatin (SS) in spontaneously hypertensive rats (SHR). Forty-five SHRs (14 weeks old, male) were randomly assigned into 3 groups (n=15):SHR group, Benazepril group (which was given benazepril 0.90 mg·kg-1·d-1) and Amlodipine group (SHRs were given amlodipine 0.45 mg· kg-1·d-1), taking WistarKyoto(WKY) as normal control (n=15), meanwhile, rats in SHR group and WKY group were given the same volume of distilled water. After 8 weeks of intervention, the expression of protein and mRNA of PZ in duodenum and SS in sinuses ventriculi was detected by enzyme-linked immunoassay and RT-PCR. After 8 weeks of intervention, compared with the WKY group, the expression of protein and mRNA of PZ in duodenum and SS in sinuses ventriculi was increased significantly in SHR group (P<0. 05). Compared with SHR group, the expression of PZ in duodenum and SS in sinuses ventriculi was decreased significantly in Benazepril group and Amlodipine group (P<0.05). Compared with Benazepril group, in Amlodipine group the expression of PZ mRNA in duodenum and SS mRNA in sinuses ventriculi was decreased more significantly (P<0.05). The regulation disorder of PZ in duodenum and SS in sinuses ventriculi exists in SHR. The antihypertensive effect of benazepril and amlodipine may be realized by regulating the expression of PZ and SS, while the regulation of amlodipine is more obvious than benazepril.

Effect of the Orally Active Growth Hormone Secretagogue MK-677 on Somatic Growth in Rats.

PurposeGrowth hormone secretagogues (GHSs) possess the ability to release growth hormone (GH) in the body. This study aimed to investigate the effects of MK-677, an orally active GHS, on somatic growth in rats.Materials and MethodsThe serum levels of GH were measured after oral administration of MK-677 to confirm GH stimulatory effects. Body weight, body length, tibia length, epiphyseal plate width, and serum levels of insulin-like growth factor (IGF)-I were measured after oral administration of 4 mg/kg of MK-677 for 6 weeks to investigate growth-promoting effects.ResultsOral administration of MK-677 at 4 mg/kg increased peak GH concentrations by 1.8-fold, compared to baseline. However, oral administration of MK-677 for 6 weeks did not increase body growth or serum levels of IGF-I. At 6 weeks after treatment, the GH response to MK-677 was abolished. Pituitary GH mRNA and hypothalamic GH-releasing hormone mRNA, and GH secretagogue receptor (GHSR) mRNA expression in the pituitary and hypothalamus did not differ between the control and treatment group. Somatostatin (SST) mRNA expression in the hypothalamus was markedly increased in the treatment group, whereas SST receptor (SSTR)-2 mRNA expression in the pituitary gland was decreased. Protein expression of hypothalamic GHSR, SST, and pituitary SSTR-2 showed patterns similar to those for mRNA expression.ConclusionOur results suggest that prolonged administration of MK-677 in rats does not promote growth despite the GH stimulatory effect of MK-677, which may be related to increased expression of SST in the hypothalamus. Further studies are needed to overcome the observed desensitization to GHS.

Obestatin stimulates the somatotrophic axis activity in sheep

The effects of obestatin (an anorexigenic peripheral peptide) on somatotrophic axis activity in ruminants have not yet been determined. The aim of this study was to investigate the consequence of intracerebroventricular infusions of obestatin on the activity of the somatotrophic axis in peripubertal female sheep. Animals were randomly divided into two groups: control group received intracerebroventricular infusions of the vehicle, and the obestatin group was infused with obestatin (25 µg/120 µL h−1). The series of four hourly infusions on three consecutive days were performed. The blood samples were collected on day 0 and on day 3. Immediately after the end of experiment sheep were slaughtered. Parts of the brains were fixed in situ for further immunohistochemical analysis, while the remaining brains were frozen for Real Time RT-qPCR analysis.Substantial changes in the activity of the somatotrophic axis were observed in obestatin-infused sheep. In those animals obestatin evoked an increase in growth hormone-releasing hormone (GHRH) mRNA expression and a decrease in somatostatin mRNA expression in the anterior hypothalamic area. Moreover, a decrease in somatostatin immunoreactivity in the periventricular nucleus and an increase in somatostatin immunopositive fibers in the median eminence were noted. Changes in the GHRH and somatostatin activity are associated with an increase in growth hormone (GH) gene expression and in the amount of GH immunoreactive material stored in the somatotrophic pituitary cells. Consequently, an increase in GH concentration in the peripheral blood, due to an increase in the number of pulses was observed.It was revealed that obestatin affects the somatostatin/GHRH/GH system at the level of protein synthesis, accumulation and release. It is suggested that obestatin participates in the mechanism modulating somatotrophic axis activity at the central level by stimulating GH release through suppression of somatostatin output. Thereby, it can be concluded that obestatin may be involved in the modulation of growth processes in sheep.

TSC1-mTOR signaling determines the differentiation of islet cells.

Neurogenin3-driven deletion of tuberous sclerosis complex 1 (Tsc1) activated mechanistic target of rapamycin complex 1 (mTORC1) measured by the upregulation of mTOR and S6 phosphorylation in islet cells. Neurogenin3-Tsc1-/- mice demonstrated a significant increase in average islet size and mean area of individual islet cell. Insulin mRNA and plasma insulin levels increased significantly after weaning. Glucagon mRNA and plasma levels increased in neonate followed by modest reduction in adult. Somatostatin mRNA and plasma levels markedly increased. Neurogenin3-Tsc1-/- mice fed standard chow demonstrated a significant improvement in glucose tolerance and no alteration in insulin sensitivity. In Neurogenin3-Tsc1-/- mice fed 45% high-fat diets, both glucose tolerance and insulin sensitivity were significantly impaired. Rapamycin reversed the activation of mTORC1, attenuated β cells hypertrophy and abolished the improvement of glucose tolerance. TSC1-mTORC1 signaling plays an important role in the development of pancreatic endocrine cells and in the regulation of glucose metabolism.

Altered Markers of Cortical γ-Aminobutyric Acid Neuronal Activity in Schizophrenia: Role of the NARP Gene.

In schizophrenia, working memory deficits appear to reflect abnormalities in the generation of gamma oscillations in the dorsolateral prefrontal cortex. The generation of gamma oscillations requires the phasic excitation of inhibitory parvalbumin-containing interneurons. Thus, gamma oscillations depend, in part, on the number of synaptic glutamate receptors on parvalbumin interneurons. However, little is known about the molecular factors that regulate glutamate receptor-mediated excitation of parvalbumin interneurons in schizophrenia. To quantify in individuals with schizophrenia the expression of immediate early genes (NARP, ARC, and SGK1) regulating glutamate synaptic neurotransmission. Postmortem brain specimens (n = 206) were obtained from individuals with schizophrenia, bipolar disorder, or major depressive disorder and from well-matched healthy persons (controls). For a study of brain tissue, quantitative polymerase chain reaction, in situ hybridization, or microarray analyses were used to measure transcript levels in the dorsolateral prefrontal cortex at gray matter, laminar, and cellular levels of resolutions. This study was conducted between January 1, 2013, and November 30, 2014. Expression levels for NARP, ARC, and SGK1 messenger RNA (mRNA) were compared between specimens from individuals with schizophrenia and controls. Diagnostic specificity was assessed by quantifying NARP mRNA levels in specimens from individuals with mood disorders. By quantitative polymerase chain reaction, levels of NARP mRNA were significantly lower by 25.6% in specimens from individuals with schizophrenia compared with the controls (mean [SD], 0.036 [0.018] vs 0.049 [0.015]; F1,114 = 21.0; P < .001). Levels of ARC (F1,112 = 0.93; P = .34) and SGK1 (F1,110 = 2.52; P = .12) were not significant. These findings were supported by in situ hybridization (NARP; individuals with schizophrenia vs controls: 40.1% lower [P = .003]) and microarray analyses (NARP; individuals with schizophrenia vs controls: 12.2% lower in layer 3 [P = .11] and 14.6% lower in layer 5 pyramidal cells [P = .001]). In schizophrenia specimens, NARP mRNA levels were positively correlated with GAD67 mRNA (r = 0.55; P < .001); the expression of GAD67 mRNA in parvalbumin interneurons is activity dependent. The NARP mRNA levels were also lower than healthy controls in bipolar disorder (-18.2%; F1,60 = 11.39; P = .001) and major depressive disorder (-21.7%; F1,30 = 5.36; P = .03) specimens, especially those from individuals with psychosis. In all 3 diagnostic groups, NARP mRNA levels were positively correlated (all r ≥ 0.53; all P ≤ .02) with somatostatin mRNA, the expression of which is activity dependent. Given the role of NARP in the formation of excitatory inputs to parvalbumin (and perhaps somatostatin) interneurons, our findings suggest that lower NARP mRNA expression contributes to lower excitatory drive onto parvalbumin interneurons in schizophrenia. This reduced excitatory drive may lead to lower synthesis of γ-aminobutyric acid in these interneurons, contributing to a reduced capacity to generate the gamma oscillations required for working memory.

Adolescent testosterone influences BDNF and TrkB mRNA and neurotrophin–interneuron marker relationships in mammalian frontal cortex

Late adolescence in males is a period of increased susceptibility for the onset of schizophrenia, coinciding with increased circulating testosterone. The cognitive deficits prevalent in schizophrenia may be related to unhealthy cortical interneurons, which are trophically dependent on brain derived neurotrophic factor. We investigated, under conditions of depleted (monkey and rat) and replaced (rat) testosterone over adolescence, changes in gene expression of cortical BDNF and TrkB transcripts and interneuron markers and the relationships between these mRNAs and circulating testosterone. Testosterone removal by gonadectomy reduced gene expression of some BDNF transcripts in monkey and rat frontal cortices and the BDNF mRNA reduction was prevented by testosterone replacement. In rat, testosterone replacement increased the potential for classical TrkB signalling by increasing the full length to truncated TrkB mRNA ratio, whereas in the monkey cortex, circulating testosterone was negatively correlated with the TrkB full length/truncated mRNA ratio. We did not identify changes in interneuron gene expression in monkey frontal cortex in response to gonadectomy, and in rat, we showed that only somatostatin mRNA was decreased by gonadectomy but not restored by testosterone replacement. We identified complex and possibly species-specific, relationships between BDNF/TrkB gene expression and interneuron marker gene expression that appear to be dependent on the presence of testosterone at adolescence in rat and monkey frontal cortices. Taken together, our findings suggest there are dynamic relationships between BDNF/TrkB and interneuron markers that are dependent on the presence of testosterone but that this may not be a straightforward increase in testosterone leading to changes in BDNF/TrkB that contributes to interneuron health.

Hypermethylation of BDNF and SST Genes in the Orbital Frontal Cortex of Older Individuals: A Putative Mechanism for Declining Gene Expression with Age.

Expression of brain-derived neurotrophic factor (BDNF) and somatostatin (SST) mRNAs in the brain decreases progressively and robustly with age, and lower BDNF and SST expression in the brain has been observed in many brain disorders. BDNF is known to regulate SST expression; however, the mechanisms underlying decreased expression of both genes are not understood. DNA methylation (DNAm) is an attractive candidate mechanism. To investigate the contribution of DNAm to the age-related decline in BDNF and SST expression, the Illumina Infinium HumanMethylation450 Beadchip Array was used to quantify DNAm of BDNF (26 CpG loci) and SST (9 CpG loci) in the orbital frontal cortices of postmortem brains from 22 younger (age <42 years) and 22 older individuals (age >60 years) with known age-dependent BDNF and SST expression differences. Relative to the younger individuals, 10 of the 26 CpG loci in BDNF and 8 of the 9 CpG loci in SST were significantly hypermethylated in the older individuals. DNAm in BDNF exons/promoters I, II, and IV negatively correlated with BDNF expression (r=-0.37, p<0.05; r=-0.40, p<0.05; r=-0.24, p=0.07), and DNAm in SST 5' UTR and first exon/intron negatively correlated with SST expression (r=-0.48, p<0.01; r=-0.63, p<0.001), respectively. An expanded set of BDNF- and GABA-related genes exhibited similar age-related changes in DNAm and correlation with gene expression. These results suggest that DNAm may be a proximal mechanism for decreased expression of BDNF, SST, and other BDNF- and GABA-related genes with brain aging and, by extension, for brain disorders in which their expression is decreased.

Effects of potassium diformate on the gastric function of weaning piglets

Potassium diformate (KDF), as an acidifier, has been shown to improve growth performance in pigs, but it is not yet known whether KDF regulates gastric function. Thus, the objective of the present study was to investigate the effects of dietary KDF on gastric function in weaning piglets. One hundred and eighty Landrace × Large White piglets (bodyweight = 5.80 ± 0.15 kg) were weaned at 28 days old and randomly allocated into two groups, with six pens in each group and 15 piglets in each pen. Piglets in the control group were fed the basal diet, whereas the KDF-treated group was fed the basal diet supplemented with 10 g/kg KDF. After 35 days of feeding, the KDF treatment improved the bodyweight (P = 0.034) and reduced the relative weight of stomach (P = 0.050), decreased the hydrochloric acid concentration (P = 0.016) in the gastric digesta and the pepsin activity in the gastric oxyntic mucosa (P = 0.001) and increased the lactic acid concentration (P = 0.001) in the gastric digesta. Furthermore, KDF treatment increased the level of somatostatin (SS) (P = 0.009), but did not change the concentration of gastrin (P = 0.497) and the activity of H+-K+-ATPase (P = 0.575) in the gastric oxyntic mucosa. However, KDF treatment downregulated the expression of SS mRNA in the gastric oxyntic mucosa (P = 0.031) and upregulated the mRNA expression of gastrin (P &lt; 0.001) and H+-K+-ATPase (P &lt; 0.001) in the gastric oxyntic mucosa. These results suggest that the effects of KDF on weaning piglets may be related to the regulation of gastric function gene expression.

Delta Cell Hyperplasia in Adult Goto-Kakizaki (GK/MolTac) Diabetic Rats.

Reduced beta cell mass in pancreatic islets (PI) of Goto-Kakizaki (GK) rats is frequently observed in this diabetic model, but knowledge on delta cells is scarce. Aiming to compare delta cell physiology/pathology of GK to Wistar rats, we found that delta cell number increased over time as did somatostatin mRNA and delta cells distribution in PI is different in GK rats. Subtle changes in 6-week-old GK rats were found. With maturation and aging of GK rats, disturbed cytoarchitecture occurred with irregular beta cells accompanied by delta cell hyperplasia and loss of pancreatic polypeptide (PPY) positivity. Unlike the constant glucose-stimulation index for insulin PI release in Wistar rats, this index declined with GK age, whereas for somatostatin it increased with age. A decrease of GK rat PPY serum levels was found. GK rat body weight decreased with increasing hyperglycemia. Somatostatin analog octreotide completely blocked insulin secretion, impaired proliferation at low autocrine insulin, and decreased PPY secretion and mitochondrial DNA in INS-1E cells. In conclusion, in GK rats PI, significant local delta cell hyperplasia and suspected paracrine effect of somatostatin diminish beta cell viability and contribute to the deterioration of beta cell mass. Altered PPY-secreting cells distribution amends another component of GK PI's pathophysiology.

Cortical inhibitory neuron disturbances in schizophrenia: role of the ontogenetic transcription factor Lhx6.

Disturbances in parvalbumin- and somatostatin-containing neurons, including deficits in the gamma-aminobutyric acid (GABA)-synthesizing enzyme GAD67 in the prefrontal cortex (PFC) in schizophrenia, may be related to disrupted pre- and/or postnatal development. Deficits in the transcription factor Lhx6, which regulates parvalbumin and somatostatin neuron development, are associated with GAD67 deficits in schizophrenia. Therefore, we investigated the potential pre- and postnatal roles of Lhx6 in GABA-related disturbances using qPCR and/or in situ hybridization to quantify PFC levels of (1) Lhx6 mRNA in a new cohort of schizophrenia subjects; (2) Lhx6 mRNA in monkeys across postnatal development; (3) GABA-related mRNAs in Lhx6 heterozygous (Lhx6+/−) mice, which model Lhx6 deficits in schizophrenia; and (4) Lhx6 mRNA in GAD67+/− mice, which model GAD67 deficits in schizophrenia. Lhx6 mRNA levels were lower (−15%) in schizophrenia and correlated with lower GAD67 mRNA levels. In addition, Lhx6 mRNA levels declined 24% from the perinatal to prepubertal periods then stabilized in monkeys. Finally, GAD67, parvalbumin, and somatostatin mRNAs were not altered in Lhx6+/− mice, and Lhx6 mRNA was not altered in GAD67+/− mice. These data suggest that PFC Lhx6 and GAD67 mRNA deficits are common components of GABA neuron pathology in schizophrenia. An excessive early postnatal decline in Lhx6 mRNA might contribute to Lhx6 mRNA deficits in schizophrenia. However, a partial loss of Lhx6 is not sufficient in isolation to produce deficits in GAD67 mRNA and vice versa, suggesting that the concurrence of Lhx6 and GAD67 mRNA deficits in schizophrenia may instead be the consequence of a common upstream factor.