Down regulation of the c-Fos/MAP kinase signaling pathway during learning memory processes coincides with low GnRH levels in aluminum chloride-induced Alzheimer's male rats.

Aluminum chloride (Al) is associated with Alzheimer's disease (AD) and reproductive disorders. But the relationship between gonadotropin-releasing hormone (GnRH) and c-Fos levels, the end product of MAP-kinase signaling, in AD is unknown, so we aimed to investigate this relationship. We exposed rats to Al dissolved in drinking water (10 and 50 mg/kg) for two and four weeks. The control group received only drinking water. At the end, the blood sample was collected under deep anesthesia and the brain was dissected on ice, and the testicular tissue was fixed in formalin. Amyloid beta (ßA) plaques in brain regions and the number of CA1 neurons were evaluated by Congo red staining and cresyl violet staining. Activation of neuronal nitric oxide synthase (nNOS) was studied using NADPH-diaphorase. The levels of c-Fos and testosterone receptors in the target area were examined immunohistochemically. Brain GnRH levels were determined by blotting, and serum levels of gonadotropins and steroids were measured by enzyme-linked immunosorbent assay (ELISA). All data were analyzed using analysis of variance (ANOVA) at α = 0.05 level. The accumulation of ßA plaque was observed along with a decrease in the number of CA1 pyramidal neurons and a significant decrease in the levels of c-Fos and GnRH in the brains of rats receiving Al, which was aligned with a significant decrease in serum levels of testosterone and LH. This study, for the first time, showed a link between dementia and a concomitant decrease in brain GnRH and c-Fos levels.

Ameliorating Effect of Standardized Rice Bran Supplement on Depressive-Like Behaviors in Ovariectomized Mice.

Menopausal depression, often associated with hormonal fluctuations such as decreased estrogen levels, imposes significant mental health burdens. Despite the antidepressant biological properties of standardized rice bran supplement (RBS), its impact on menopausal depression and underlying mechanisms remains largely unexplored. In this study, we investigated the antidepressant effects of RBS in a mouse model of estrogen deficiency-induced depression. Ovariectomized (OVX) mice received oral doses of RBS (250 and 1000 mg/kg) and 17ß estradiol over a 20-week period. RBS administration resulted in decreased immobility time in the tail suspension and forced swim tests, along with increased locomotor activity in the open field test. Furthermore, RBS enhanced nitric oxide production and neuronal nitric oxide synthase (nNOS) expression in the hippocampi of OVX mice. Additionally, RBS administration phosphorylated extracellular signal-regulated kinase (ERK), cAMP response element-binding protein (CREB), and tropomyosin receptor kinase B and increased the protein expression of brain-derived neurotrophic factor (BDNF) in the hippocampus. These findings suggest that RBS alleviated depressive behaviors in OVX mice by augmenting hippocampal nNOS expression and activating the ERK-CREB-BDNF signaling pathway. Therefore, based on these results, we propose that RBS is a promising agent to treat menopausal depression, a challenging condition.

Comparison of the Influence of Bisphenol A and Bisphenol S on the Enteric Nervous System of the Mouse Jejunum.

Bisphenols are dangerous endocrine disruptors that pollute the environment. Due to their chemical properties, they are globally used to produce plastics. Structural similarities to oestrogen allow bisphenols to bind to oestrogen receptors and affect internal body systems. Most commonly used in the plastic industry is bisphenol A (BPA), which also has negative effects on the nervous, immune, endocrine, and cardiovascular systems. A popular analogue of BPA-bisphenol S (BPS) also seems to have harmful effects similar to BPA on living organisms. Therefore, with the use of double immunofluorescence labelling, this study aimed to compare the effect of BPA and BPS on the enteric nervous system (ENS) in mouse jejunum. The study showed that both studied toxins impact the number of nerve cells immunoreactive to substance P (SP), galanin (GAL), vasoactive intestinal polypeptide (VIP), the neuronal isoform of nitric oxide synthase (nNOS), and vesicular acetylcholine transporter (VAChT). The observed changes were similar in the case of both tested bisphenols. However, the influence of BPA showed stronger changes in neurochemical coding. The results also showed that long-term exposure to BPS significantly affects the ENS.

Oral Exposure to Microplastics Affects the Neurochemical Plasticity of Reactive Neurons in the Porcine Jejunum.

Plastics are present in almost every aspect of our lives. Polyethylene terephthalate (PET) is commonly used in the food industry. Microparticles can contaminate food and drinks, posing a threat to consumers. The presented study aims to determine the effect of microparticles of PET on the population of neurons positive for selected neurotransmitters in the enteric nervous system of the jejunum and histological structure. An amount of 15 pigs were divided into three groups (control, receiving 0.1 g, and 1 g/day/animal orally). After 28 days, fragments of the jejunum were collected for immunofluorescence and histological examination. The obtained results show that histological changes (injury of the apical parts of the villi, accumulations of cellular debris and mucus, eosinophil infiltration, and hyperaemia) were more pronounced in pigs receiving a higher dose of microparticles. The effect on neuronal nitric oxide synthase-, and substance P-positive neurons, depends on the examined plexus and the dose of microparticles. An increase in the percentage of galanin-positive neurons and a decrease in cocaine and amphetamine-regulated transcript-, vesicular acetylcholine transporter-, and vasoactive intestinal peptide-positive neurons do not show such relationships. The present study shows that microparticles can potentially have neurotoxic and pro-inflammatory effects, but there is a need for further research to determine the mechanism of this process and possible further effects.

The human major sublingual gland and its neuropeptidergic and nitrergic innervations.

BACKGROUND: What textbooks usually call the sublingual gland in humans is in reality a tissue mass of two types of salivary glands, the anteriorly located consisting of a cluster of minor sublingual glands and the posteriorly located major sublingual gland with its outlet via Bartholin's duct. Only recently, the adrenergic and cholinergic innervations of the major sublingual gland was reported, while information regarding the neuropeptidergic and nitrergic innervations is still lacking. METHODS: Bioptic and autoptic specimens of the human major sublingual gland were examined by means of immunohistochemistry for the presence of vasoactive intestinal peptide (VIP)-, neuropeptide Y (NPY)-, substance P (SP)-, calcitonin gene related-peptide (CGRP)-, and neuronal nitric oxide synthase (nNOS)-labeled neuronal structures. RESULTS: As to the neuropeptidergic innervation of secretory cells (here in the form of mucous tubular and seromucous cells), the findings showed many VIP-containing nerves, few NPY- and SP-containing nerves and a lack of CGRP-labeled nerves. As to the neuropeptidergic innervation of vessels, the number of VIP-containing nerves was modest, while, of the other neuropeptide-containing nerves under study, only few (SP and CGRP) to very few (NPY) nerves were observed. As to the nitrergic innervation, nNOS-containing nerves were very few close to secretory cells and even absent around vessels. CONCLUSION: The various innervation patterns may suggest potential transmission mechanisms involved in secretory and vascular responses of the major sublingual gland.

The vascular footprint in cardiac homeostasis and hypertensive heart disease-A link between apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase.

Recent studies have suggested a connection between disturbances of the apelin system and various cardiac pathologies, including hypertension, heart failure, and atherosclerosis. Vascular endothelial growth factor is crucial for cardiac homeostasis as a critical molecule in cardiac angiogenesis. Neuronal nitric oxide synthase is an essential enzyme producing nitric oxide, a key regulator of vascular tone. The present study aims to shed light upon the complex interactions between these three vital signaling molecules and examine their changes with the progression of hypertensive heart disease. We used two groups of spontaneously hypertensive rats and age-matched Wistar rats as controls. The expression of the apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase were assessed immunohistochemically. We used capillary density and cross-sectional area of the cardiomyocytes as quantitative parameters of cardiac hypertrophy. Immunoreactivity of the molecules was more potent in both ventricles of spontaneously hypertensive rats compared with age-matched controls. However, capillary density was lower in both ventricles of the two age groups of spontaneously hypertensive rats compared with controls, and the difference was statistically significant. In addition, the cross-sectional area of the cardiomyocytes was higher in both ventricles of the two age groups of spontaneously hypertensive rats compared with controls, and the difference was statistically significant. Our study suggests a potential link between the apelin receptor, vascular endothelial growth factor, and neuronal nitric oxide synthase in cardiac homeostasis and the hypertensive myocardium. Nevertheless, further research is required to better comprehend these interactions and their potential therapeutic implications.

Cannabidiol induces systemic analgesia through activation of the PI3Kγ/nNOS/NO/KATP signaling pathway in neuropathic mice. A KATP channel S-nitrosylation-dependent mechanism.

BACKGROUND: Cannabidiol (CBD) is the second most abundant pharmacologically active component present in Cannabis sp. Unlike Δ-9-tetrahydrocannabinol (THC), it has no psychotomimetic effects and has recently received significant interest from the scientific community due to its potential to treat anxiety and epilepsy. CBD has excellent anti-inflammatory potential and can be used to treat some types of inflammatory and neuropathic pain. In this context, the present study aimed to evaluate the analgesic mechanism of cannabidiol administered systemically for the treatment of neuropathic pain and determine the endogenous mechanisms involved with this analgesia. METHODS: Neuropathic pain was induced by sciatic nerve constriction surgery, and the nociceptive threshold was measured using the paw compression test in mice. RESULTS: CBD produced dose-dependent antinociception after intraperitoneal injection. Selective inhibition of PI3Kγ dose-dependently reversed CBD-induced antinociception. Selective inhibition of nNOS enzymes reversed the antinociception induced by CBD, while selective inhibition of iNOS and eNOS did not alter this antinociception. However, the inhibition of cGMP production by guanylyl cyclase did not alter CBD-mediated antinociception, but selective blockade of ATP-sensitive K+ channels dose-dependently reversed CBD-induced antinociception. Inhibition of S-nitrosylation dose-dependently and completely reversed CBD-mediated antinociception. CONCLUSION: Cannabidiol has an antinociceptive effect when administered systemically and this effect is mediated by the activation of PI3Kγ as well as by nitric oxide and subsequent direct S-nitrosylation of KATP channels on peripheral nociceptors.

Restoration of nNOS Expression Rescues Autistic-Like Phenotypes Through Normalization of AMPA Receptor-Mediated Neurotransmission.

Autism spectrum disorder (ASD) is associated with a range of abnormalities characterized by deficits in socialization, communication, repetitive behaviors, and restricted interests. We have recently shown that neuronal nitric oxide synthase (nNOS) expression was decreased in the basolateral amygdala (BLA) of mice after postnatal valproic acid exposure. Neuronal activity-regulated pentraxin (Narp) could contribute to the regulation of the GluA4 2-amino-3-(5-methyl-3-oxo-1,2-oxazol-4-yl) propanoic acid (AMPA) subunits which are predominantly expressed in interneurons. However, the specific role of nNOS re-expression on excitatory neurotransmitter with relevance to ASD core symptoms in VPA-treated animals remains to be elucidated. Herein, nNOS overexpression using a lentiviral vector and L-arginine-activating PI3K-Akt-mTOR signaling can restore nNOS expression in the BLA induced by VPA. Restoration of nNOS expression in these mice was sufficient to reduce the severity of ASD-like behavioral patterns such that animals exhibited decreases in abnormal social interactions and communication, stereotyped/repetitive behaviors, and anxiety-like traits. Most strikingly, re-expression of nNOS upregulated surface expression of Narp and GluA4 in nNOS-positive interneuron as shown by immunoprecipitation and Western blotting. Whole-cell patch-clamp recordings demonstrated that restoration of nNOS had a significant enhancing effect on AMPA receptor-mediated excitatory glutamatergic synaptic neurotransmission, which was inhibited by disturbing the interaction between Narp and GluA4 in acutely dissociated BLA slices. Overall, these data offer a scientific basis for the additional study of nNOS re-expression as a promising therapeutic target by correcting AMPA receptor-mediated synaptic function in ASD and related neurodevelopmental disorders.

The Physiological Function of nNOS-Associated CAPON Proteins and the Roles of CAPON in Diseases.

In this review, the structure, isoform, and physiological role of the carboxy-terminal PDZ ligand of neuronal nitric oxide synthase (CAPON) are summarized. There are three isoforms of CAPON in humans, including long CAPON protein (CAPON-L), short CAPON protein (CAPON-S), and CAPON-S' protein. CAPON-L includes three functional regions: a C-terminal PDZ-binding motif, carboxypeptidase (CPE)-binding region, and N-terminal phosphotyrosine (PTB) structural domain. Both CAPON-S and CAPON-S' only contain the C-terminal PDZ-binding motif. The C-terminal PDZ-binding motif of CAPON can bind with neuronal nitric oxide synthase (nNOS) and participates in regulating NO production and neuronal development. An overview is given on the relationship between CAPON and heart diseases, diabetes, psychiatric disorders, and tumors. This review will clarify future research directions on the signal pathways related to CAPON, which will be helpful for studying the regulatory mechanism of CAPON. CAPON may be used as a drug target, which will provide new ideas and solutions for treating human diseases.

Nitric Oxide Synthase Inhibition Prevents Cell Proliferation in Glioblastoma.

Glioblastoma multiforme (GBM) is a prevalent and aggressive primary brain tumor, presenting substantial treatment challenges and high relapse rates. GBM is characterized by alterations in molecular signaling and enzyme expression within malignant cells. This tumor exhibits elevated nitric oxide (NO.) levels. NO. is a crucial signaling molecule involved in the regulation of neuronal functions, synaptic transmission, and cell proliferation. It is primarily synthesized from L-arginine by nitric oxide synthase (NOS) enzymes. The increased levels of NO. in GBM stem from dysregulated activity and expression of clinically relevant NOS isoforms, particularly inducible NOS (iNOS) and neuronal NOS (nNOS). Based on this knowledge, we hypothesize that targeted pharmacological intervention with N6-(1-iminoethyl)-L-lysine (L-NIL), an iNOS inhibitor, and 7-Nitroindazole (7-NI), an nNOS inhibitor, may suggest a promising therapeutic strategy for the treatment of GBM. To test our hypothesis, we utilized the U87-MG cell line as an in vitro model of GBM. Our results showed that treatment with L-NIL and 7-NI led to a reduction in NO. levels, NOS activity, and clonogenic proliferation in U87-MG cells. These findings suggest that NO. and NOS enzymes might be prospective therapeutic targets for GBM.

ARL-17477 is a dual inhibitor of NOS1 and the autophagic-lysosomal system that prevents tumor growth in vitro and in vivo.

ARL-17477 is a selective neuronal nitric oxide synthase (NOS1) inhibitor that has been used in many preclinical studies since its initial discovery in the 1990s. In the present study, we demonstrate that ARL-17477 exhibits a NOS1-independent pharmacological activity that involves inhibition of the autophagy-lysosomal system and prevents cancer growth in vitro and in vivo. Initially, we screened a chemical compound library for potential anticancer agents, and identified ARL-17477 with micromolar anticancer activity against a wide spectrum of cancers, preferentially affecting cancer stem-like cells and KRAS-mutant cancer cells. Interestingly, ARL-17477 also affected NOS1-knockout cells, suggesting the existence of a NOS1-independent anticancer mechanism. Analysis of cell signals and death markers revealed that LC3B-II, p62, and GABARAP-II protein levels were significantly increased by ARL-17477. Furthermore, ARL-17477 had a chemical structure similar to that of chloroquine, suggesting the inhibition of autophagic flux at the level of lysosomal fusion as an underlying anticancer mechanism. Consistently, ARL-17477 induced lysosomal membrane permeabilization, impaired protein aggregate clearance, and activated transcription factor EB and lysosomal biogenesis. Furthermore, in vivo ARL-17477 inhibited the tumor growth of KRAS-mutant cancer. Thus, ARL-17477 is a dual inhibitor of NOS1 and the autophagy-lysosomal system that could potentially be used as a cancer therapeutic.

Neuroprotective Effects of Nanowired Delivery of Cerebrolysin with Mesenchymal Stem Cells and Monoclonal Antibodies to Neuronal Nitric Oxide Synthase in Brain Pathology Following Alzheimer's Disease Exacerbated by Concussive Head Injury.

Concussive head injury (CHI) is one of the major risk factors in developing Alzheimer's disease (AD) in military personnel at later stages of life. Breakdown of the blood-brain barrier (BBB) in CHI leads to extravasation of plasma amyloid beta protein (ΑßP) into the brain fluid compartments precipitating AD brain pathology. Oxidative stress in CHI or AD is likely to enhance production of nitric oxide indicating a role of its synthesizing enzyme neuronal nitric oxide synthase (NOS) in brain pathology. Thus, exploration of the novel roles of nanomedicine in AD or CHI reducing NOS upregulation for neuroprotection are emerging. Recent research shows that stem cells and neurotrophic factors play key roles in CHI-induced aggravation of AD brain pathologies. Previous studies in our laboratory demonstrated that CHI exacerbates AD brain pathology in model experiments. Accordingly, it is quite likely that nanodelivery of NOS antibodies together with cerebrolysin and mesenchymal stem cells (MSCs) will induce superior neuroprotection in AD associated with CHI. In this review, co-administration of TiO2 nanowired cerebrolysin - a balanced composition of several neurotrophic factors and active peptide fragments, together with MSCs and monoclonal antibodies (mAb) to neuronal NOS is investigated for superior neuroprotection following exacerbation of brain pathology in AD exacerbated by CHI based on our own investigations. Our observations show that nanowired delivery of cerebrolysin, MSCs and neuronal NOS in combination induces superior neuroprotective in brain pathology in AD exacerbated by CHI, not reported earlier.

Calcium transients in nNOS neurons underlie distinct phases of the neurovascular response to barrel cortex activation in awake mice.

Neuronal nitric oxide (NO) synthase (nNOS), a Ca2+ dependent enzyme, is expressed by distinct populations of neocortical neurons. Although neuronal NO is well known to contribute to the blood flow increase evoked by neural activity, the relationships between nNOS neurons activity and vascular responses in the awake state remain unclear. We imaged the barrel cortex in awake, head-fixed mice through a chronically implanted cranial window. The Ca2+ indicator GCaMP7f was expressed selectively in nNOS neurons using adenoviral gene transfer in nNOScre mice. Air-puffs directed at the contralateral whiskers or spontaneous motion induced Ca2+ transients in 30.2 ± 2.2% or 51.6 ± 3.3% of nNOS neurons, respectively, and evoked local arteriolar dilation. The greatest dilatation (14.8 ± 1.1%) occurred when whisking and motion occurred simultaneously. Ca2+ transients in individual nNOS neurons and local arteriolar dilation showed various degrees of correlation, which was strongest when the activity of whole nNOS neuron ensemble was examined. We also found that some nNOS neurons became active immediately prior to arteriolar dilation, while others were activated gradually after arteriolar dilatation. Discrete nNOS neuron subsets may contribute either to the initiation or to the maintenance of the vascular response, suggesting a previously unappreciated temporal specificity to the role of NO in neurovascular coupling.

Male minipuberty involves the gonad-independent activation of preoptic nNOS neurons.

BACKGROUND: The maturation of the hypothalamic-pituitary-gonadal (HPG) axis is crucial for the establishment of reproductive function. In female mice, neuronal nitric oxide synthase (nNOS) activity appears to be key for the first postnatal activation of the neural network promoting the release of gonadotropin-releasing hormone (GnRH), i.e. minipuberty. However, in males, the profile of minipuberty as well as the role of nNOS-expressing neurons remain unexplored. METHODS: nNOS-deficient and wild-type mice were studied during postnatal development. The expression of androgen (AR) and estrogen receptor alpha (ERα) as well as nNOS phosphorylation were evaluated by immunohistochemistry in nNOS neurons in the median preoptic nucleus (MePO), where most GnRH neuronal cell bodies reside, and the hormonal profile of nNOS-deficient male mice was assessed using previously established radioimmunoassay and ELISA methods. Gonadectomy and pharmacological manipulation of ERα were used to elucidate the mechanism of minipubertal nNOS activation and the maturation of the HPG axis. RESULTS: In male mice, minipubertal FSH release occurred at P23, preceding the LH surge at P30, when balanopreputial separation occurs. Progesterone and testosterone remained low during minipuberty, increasing around puberty, whereas estrogen levels were high throughout postnatal development. nNOS neurons showed a sharp increase in Ser1412 phosphorylation of nNOS at P23, a phenomenon that occurred even in the absence of the gonads. In male mice, nNOS neurons did not appear to express AR, but abundantly expressed ERα throughout postnatal development. Selective pharmacological blockade of ERα during the infantile period blunted Ser1412 phosphorylation of nNOS at P23. CONCLUSIONS: Our results show that the timing of minipuberty differs in male mice when compared to females, but as in the latter, nNOS activity in the preoptic region plays a role in this process. Additionally, akin to male non-human primates, the profile of minipuberty in male mice is shaped by sex-independent mechanisms, and possibly involves extragonadal estrogen sources.

Adrenomedullin in paraventricular nucleus attenuates adipose afferent reflex and sympathoexcitation via receptors mediated nitric oxide-gamma-aminobutyric acid A type receptor pathway in rats with obesity-related hypertension.

BACKGROUND: Hypothalamic paraventricular nucleus (PVN) is an important central site for the control of the adipose afferent reflex (AAR) that increases sympathetic outflow and blood pressure in obesity-related hypertension (OH). METHOD: In this study, we investigated the effects of nitric oxide (NO) and cardiovascular bioactive polypeptide adrenomedullin (ADM) in the PVN on AAR and sympathetic nerve activity (SNA) in OH rats induced by a high-fat diet. RESULTS: The results showed that ADM, total neuronal NO synthase (nNOS) and phosphorylated-nNOS protein expression levels in the PVN of the OH rats were down-regulated compared to the control rats. The enhanced AAR in OH rats was attenuated by PVN acute application of NO donor sodium nitroprusside (SNP), but was strengthened by the nNOS inhibitor nNOS-I, guanylyl cyclase inhibitor (1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one, ODQ) and gamma-aminobutyric acid A type receptor (GABAA) antagonist Bicuculline. Moreover, PVN ADM microinjection not only decreased basal SNA but also attenuated the enhanced AAR in OH rats, which were effectively inhibited by ADM receptor antagonist ADM22-52, nNOS-I, ODQ or Bicuculline pretreatment. Bilateral PVN acute microinjection of ADM also caused greater increases in NO and cyclic guanosine monophosphate (cGMP) levels, and nNOS phosphorylation. Adeno-associated virus vectors encoding ADM (AAV-ADM) transfection in the PVN of OH rats not only decreased the elevated AAR, basal SNA and blood pressure (BP), but also increased the expression and activation of nNOS. Furthermore, AAV-ADM transfection improved vascular remodeling in OH rats. CONCLUSION: Taken together, our data highlight the roles of ADM in improving sympathetic overactivation, enhanced AAR and hypertension, and its related mechanisms associated with receptors mediated NO-cGMP-GABAA pathway in OH condition.

Postnatal development of the enteric neurons expressing neuronal nitric oxide synthase.

Neurons, expressing neuronal nitric oxide synthase (nNOS) in the enteric ganglia are inhibitory motor neurons or interneurons. The aim of the study was to identify the percentage, cross-sectional area of nNOS-immunoreactive (IR) neurons and their colocalization with choline acetyltransferase (ChAT), vasoactive intestinal polypeptide (VIP), and neuropeptide Y in the intramural ganglia of the myenteric (MP) and submucous plexus (SP) of the small intestine (SI) and large intestine (LI) of rats of different age groups using immunohistochemical methods. In the intramural ganglia of the MP, the largest percentage of nNOS-IR neurons was detected in newborn rats in the LI (81 ± 0.9%) and SI (48 ± 4.1%). Subsequently, it decreased in ontogenesis up to 60 days of life (26 ± 0.9% LI, 29 ± 3.2% SI), and did not change until senescence. In the SP, abundant nNOS-IR neurons were also detected in newborns (82 ± 7.0% SI, 85 ± 3.2% LI), while their percentage decreased significantly in the next 20 days. Furthermore, a very small number of nNOS-IR neurons was detected in 30-day- and 2-month-old animals, but they again appeared in large numbers in aged rats. In the MP, the highest percentage of nNOS+/ChAT+ neurons was in 1-day-old, 10-day-old, and 2-year-old rats. In the SP, the largest number of nNOS-IR neurons colocalized ChAT regardless of age. In the MP of all rats, many nNOS-IR neurons colocalized VIP, and the maximal percentage of nNOS+/VIP+ neurons was found in 2-year-old rats, minimal-in newborns. In conclusion, nNOS expression in neurons of the gut is decreased in early postnatal ontogenesis and subsequently increased in aged rats.

Neuronal Nitric Oxide Synthase as a Shared Target for the Effects of Adiponectin and Resistin on the Mechanical Responses of the Mouse Gastric Fundus.

It has been reported that adiponectin (ADPN) and resistin are co-secreted by white mouse adipocytes and exert similar inhibitory effects in the mouse gastric fundus, in which resistin was observed to increase neuronal nitric oxide synthase (nNOS) expression. On these grounds, the present work aimed to investigate whether the effects of the two adipokines on the neurally-induced relaxant responses potentiate each other and whether there is a possible correlation with changes in nNOS expression in preparations from the mouse gastric fundus. In carbachol (CCh)-precontracted strips, electrical field stimulation elicited nitrergic relaxant responses, whose amplitude was increased by ADPN or resistin, but no additional enhancements were observed in their concomitant presence. Western blot and immunofluorescence analyses revealed that ADPN, like resistin, was able to up-regulate nNOS expression and to increase the percentage of nNOS-positive neurons in the myenteric plexus: co-treatment with the two adipokines did not induce additional changes. The results indicate that the two adipokines modulate nitrergic neurotransmission, and both do so by up-regulating nNOS expression. Therefore, nNOS appears to be a shared target for the two adipokines' effects, which, rather than mutually reinforcing each other, may represent a dual physiological control mechanism to guarantee gastric fundus relaxation.

The interleukin-enhanced binding factor 3-mediated inhibition of nitric oxide production via phosphoinositide 3-kinase/protein kinase B pathway contributes to central cardiovascular regulation in the rostral ventrolateral medulla in hypertension.

Hypertension is characterized by sympathetic hyperactivity, which is related to the overexcitation of the presympathetic neurons in the rostral ventrolateral medulla (RVLM). Nitric oxide (NO) has been reported to be a vital neuromodulator involved in central cardiovascular regulation. However, the mechanism of interleukin-enhanced binding factor 3 (ILF3) participating in blood pressure (BP) regulation is still unclear. Therefore, this study aims to clarify the role of ILF3 within the rostral ventrolateral medulla (RVLM) in regulating NO in hypertension. It was found that the expression level of ILF3 was significantly increased in the RVLM of spontaneously hypertensive rats (SHR) compared with Wistar-Kyoto (WKY) rats through microarray gene expression analysis, Western blot, and immunofluorescence. Overexpression of ILF3 by injecting constructed adenovirus into the RVLM increased the BP and renal sympathetic nerve activity (RSNA) of the WKY rats, significantly decreasing NO production and neuronal nitric oxide synthase (nNOS) expression. Knockdown of ILF3 in the RVLM of SHR significantly reduced BP but increased NO production and the neuronal nitric oxide synthase (nNOS) expression. Furthermore, it was found that the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway was activated via Western blotting in the RVLM after overexpression of ILF3, whereas it was attenuated after knockdown of ILF3 in SHR. In addition, inhibition of PI3K by intracisternal infusion of the PI-103 attenuated the increase in Akt phosphorylation and decrease in nNOS expression and NO production caused by overexpressing ILF3, which ultimately blunted high BP induced by overexpressing ILF3. Taken together, this current study suggests that ILF3 participates in high BP via reducing NO production in the RVLM through PI3K/Akt pathway.

Nitric Oxide Linked to mGluR5 Upregulates BDNF Synthesis by Activating MMP2 in the Caudate and Putamen after Challenge Exposure to Nicotine in Rats.

Nitric oxide (NO) linked to glutamate receptors in the caudate and putamen (CPu) regulates neuroadaptation after drug exposure. Matrix-metalloproteinase (MMP), a Ca2+-dependent zinc-containing endopeptidase, increases mature brain-derived neurotrophic factor (BDNF) synthesis after drug exposure in the brain. The present study determined that NO synthesis linked to metabotropic glutamate receptor subtype 5 (mGluR5) stimulation after challenge exposure to nicotine activates MMP, which upregulates BDNF synthesis in the CPu. Subcutaneous injection of challenge nicotine (1.0 mg/kg) after repeated injections of nicotine (1.0 mg/kg/day) for 14 days and 7 days of nicotine withdrawal increased MMP2 activity and BDNF expression in the CPu of rats. These increases were prevented by the bilateral intra-CPu infusion of the mGluR5 antagonist, MPEP (0.1 nmol/side), the IP3 receptor antagonist, xestospongin C (0.004 nmol/side) or the neuronal nitric oxide synthase (nNOS) and NO inhibitor, Nω-propyl (0.1 nmol/side) prior to the challenge nicotine. Furthermore, bilateral intra-CPu infusion of the MMP2 inhibitor, OA-Hy (1 nmol/side) prevented the challenge nicotine-induced increase in the expression of BDNF. These findings suggest that elevation of NO synthesis linked to mGluR5 potentiates BDNF synthesis via activation of MMP2 after challenge exposure to nicotine in the CPu of rats.

Co-administration of sodium hydrosulfide and tadalafil modulates hypoxia and oxidative stress on bladder dysfunction in a rat model of bladder outlet obstruction.

PURPOSE: This study aimed to assess the possible healing effect of combination treatment with a hydrogen sulfide (H2S) donor, sodium hydrosulfide (NaHS) plus tadalafil on partial bladder outlet obstruction (PBOO)-induced bladder dysfunction. MATERIALS AND METHODS: A total of 75 male Sprague-Dawley rats aged 10-wk and 300-350g were divided into five groups; control; PBOO; PBOO+NaHS (5.6mg/kg/day, i.p., 6-wk); PBOO+tadalafil (2mg/kg/day, oral, 6-wk) and PBOO+NaHS+tadalafil. PBOO was created by partial urethral ligation. 6 weeks after obstruction, the in vitro contractile responses of the detrusor muscle and Western blotting, H2S and malondialdehyde assay were performed in bladder tissues. RESULTS: There was an increase in bladder weight(p<0.001) and a decrease in contractile responses to KCL(p<0.001), carbachol(p<0.01), electrical field stimulation(p<0.05) and ATP (p<0.001) in the detrusor smooth muscle of obstructed rats which was normalized after the combination treatment. Cystathionine γ-lyase and cystathionine ß-synthase, and nuclear factor kappa B protein levels did not significantly differ among groups. The obstruction induced decrement in 3-mercaptopyruvate sulfur transferase protein expression(p<0.001) and H2S levels(p<0.01) as well as increment in protein expressions of neuronal nitric oxide synthase (NO, p<0.001), endothelial NOS (p<0.05), inducible NOS(p<0.001), hypoxia-inducible factor 1-alpha (p<0.01), and malondialdehyde levels (p<0.01), when combined treatment entirely normalized. CONCLUSIONS: Combination therapy has beneficial effects on bladder dysfunction via regulating both H2S and nitric oxide pathways as well as downregulation of oxidative stress and hypoxia. The synergistic effect of H2S and nitric oxide is likely to modulate bladder function, which supports the combined therapy for enhancing clinical outcomes in men with BPH/LUTS.