Naoya Hashikawa

Candesartan Cilexetil Improves Angiotensin II Type 2 Receptor–Mediated Neurite Outgrowth via the PI3K-Akt Pathway in Fructose-Induced Insulin-Resistant Rats

We have shown previously that stimulation of the angiotensin II type 2 receptor (AT2R) results in nerve facilitation. In this study, we determined the capacity of candesartan to correct expression patterns characteristic of neuropathy and AT2R-mediated neurite outgrowth in the fructose-induced insulin-resistant rat, which is one of the human hyperinsulinemia models. Wistar rats received a 15% (w/v) fructose solution in their drinking water for 4 weeks (fructose-drinking rats [FDRs]), with or without candesartan (5 mg/kg/day). We evaluated physiological and behavioral parameters and performed immunohistochemical studies. We found that the FDR developed insulin resistance and downregulated both AT2R neuronal function and phosphorylated Akt expression in dorsal root ganglia (DRG) neurons. Candesartan improved neurite outgrowth in the FDR, which was associated with the restoration of AT2R and phosphorylated Akt expression. Furthermore, downregulation of phosphoinositide 3-kinase (PI3K) inhibited AT2R-mediated neurite outgrowth in control DRG cells. PI3K activation increased AT2R-mediated neurite outgrowth and phosphorylated Akt expression in FDR DRG cells. These results suggest that the decrease of AT2R-mediated neurite outgrowth in FDRs is likely to be the result of decreased PI3K-dependent Akt activation. Candesartan improved AT2R neuronal function and Akt phosphorylation, which were associated with sensory nerve defects and insulin sensitivity in the FDR.

Hyperinsulinemia induces hypertension associated with neurogenic vascular dysfunction resulting from abnormal perivascular innervations in rat mesenteric resistance arteries

We previously reported that chronic hyperinsulinemia and insulin resistance induced by fructose-drinking loading elicited hypertension associated with abnormal neuronal regulation of vascular tone in an in vivo study using pithed rats. Therefore, to further clarify the detailed mechanisms of perivascular nervous system malfunction induced by chronic hyperinsulinemia, we investigated the neurogenic vascular responses and distribution of perivascular nerves using mesenteric vascular beds isolated from fructose-loaded rats with hyperinsulinemia. Male Wistar rats (6 weeks old) received 15% fructose solution as drinking fluid for 10 weeks (fructose-drinking rats, FDR), which resulted in significant increases in plasma levels of insulin, the glucose-insulin index, blood norepinephrine (NE) levels and systolic blood pressure, but not blood glucose levels, when compared with normal water-drinking rats (control rats). In perfused mesenteric vascular beds of FDR, enhanced adrenergic nerve-mediated vasoconstriction with no effect on NE-induced vasoconstriction and decreased calcitonin gene-related peptide (CGRP)-containing nerve-mediated vasodilation with no effect on CGRP-induced vasodilation were observed. Immunohistochemistry studies showed increased density of neuropeptide Y immunopositive adrenergic fibers and reduced density of CGRP immunopositive fibers in mesenteric arteries of FDR. Furthermore, FDR showed decreased CGRP content in dorsal root ganglia. These findings suggest that dysfunction of the neuronal vascular control system resulting from abnormal innervation of mesenteric perivascular nerves induced by the hyperinsulinemic state is responsible for the development of hypertension in FDR.

Calcitonin gene-related peptide pre-administration acts as a novel antidepressant in stressed mice.

Calcitonin gene-related peptide (CGRP) is a neuropeptide that has potent vasodilator properties and is involved in various behavioral disorders. The relationship between CGRP and depression-like behavior is unclear. In this study, we used chronically stressed mice to investigate whether CGRP is involved in depression-like behavior. Each mouse was exposed to restraint and water immersion stress for 15 days. After stress exposure, mice were assessed using behavioral tests: open field test, forced swim test and sucrose preference test. Serum corticosterone levels, hippocampal proliferation and mRNA expression of neurotrophins were measured. After stress exposure, mice exhibited depression-like behavior and decreased CGRP mRNA levels in the hippocampus. Although intracerebroventricular CGRP administration (0.5 nmol) did not alter depression-like behavior after 15-day stress exposure, a single CGRP administration into the brain, before the beginning of the 15-day stress exposure, normalized the behavioral dysfunctions and increased nerve growth factor (Ngf) mRNA levels in stressed mice. Furthermore, in the mouse E14 hippocampal cell line, CGRP treatment induced increased expression of Ngf mRNA. The NGF receptor inhibitor K252a inhibited CGRP’s antidepressant-like effects in stressed mice. These results suggest that CGRP expression in the mouse hippocampus is associated with depression-like behavior and changes in Ngf mRNA levels.

The Akt-nitric oxide-cGMP pathway contributes to nerve growth factor-mediated neurite outgrowth in apolipoprotein E knockout mice

Apolipoprotein E (apo)-deficient [apoE(−/−)] mice have peripheral sensory nerve defects and a reduced and delayed response to noxious thermal stimuli. However, to date, no report has focused on the influence of apoE deficiency on calcitonin gene-related peptide (CGRP)-containing nerve fiber extensions. We have shown that the density of CGRP-containing nerve fibers decreases in mesenteric arteries of apoE(−/−) mice compared with wild-type mice. Here, we investigated whether apoE deficiency is involved in nerve growth factor (NGF)-induced CGRP-containing nerve regeneration using apoE(−/−) mice. NGF-mediated CGRP-like immunoreactivity (LI)-neurite outgrowth in apoE(−/−) cultured dorsal root ganglia (DRG) cells was significantly lower than that in wild-type cultures. However, the level of NGF receptor mRNA in apoE(−/−) DRG cells was similar to that in wild-type mice. To clarify the mechanism of the impaired ability of NGF-mediated neurite outgrowth, we focused on the Akt-nitric oxide (NO)-cGMP pathway. Expression of phosphorylated Akt was significantly reduced in apoE(−/−) DRG. The NO donor, sodium nitroprusside or S-nitroso-N-acetylpenicillamine, did not affect NGF-mediated neurite outgrowth in apoE(−/−) cultured DRG cells. However, 8-bromoguanosine 3′,5′-cyclic monophosphate sodium salt n-hydrate, a cGMP analog, induced NGF-mediated nerve facilitation similar to wild-type NGF-mediated neurite outgrowth levels. Furthermore, in apoE(−/−) DRG, soluble guanylate cyclase expression was significantly lower than that in wild-type DRG. These results suggest that in apoE(−/−) mice the Akt-NO-cGMP pathway is impaired, which may be caused by NGF-mediated CGRP-LI-neurite outgrowth defects.

HSP105 prevents depression-like behavior by increasing hippocampal brain-derived neurotrophic factor levels in mice

The gastric ulcer drug teprenone has central nervous system effects, specifically reducing depression-like behavior in mice. Heat shock proteins (HSPs) are stress-induced chaperones that are involved in neurological disease. Although increasingly implicated in behavioral disorders, the mechanisms of HSP action, and the relevant functional pathways, are still unclear. We examined whether oral administration of geranylgeranylacetone (GGA), a known HSP inducer, produced an antidepressant effect in a social defeat stress model of depression in mice. We also investigated the possible molecular mechanisms involved, particularly focusing on hippocampal neurogenesis and neurotrophic factor expression. In stressed mice, hippocampal HSP105 expression decreased. However, administration of GGA increased HSP105 expression and improved depression-like behavior, induced hippocampal cell proliferation, and elevated brain-derived neurotrophic factor (BDNF) levels in mouse hippocampus. Co-treatment with GGA and the BDNF receptor inhibitor K252a suppressed the antidepressant effects of GGA. HSP105 knockdown decreased BDNF mRNA levels in HT22 hippocampal cell lines and hippocampal tissue and inhibited the GGA-mediated antidepressant effect. These observations suggest that GGA administration is a therapeutic candidate for depressive diseases by increasing hippocampal BDNF levels via HSP105 expression.

Angiotensin II AT2 receptors regulate NGF-mediated neurite outgrowth via the NO-cGMP pathway

We investigated whether Angiotensin II type 2 (AT2) receptor activation was involved in NGF-induced nerve regeneration. NGF-mediated neurite outgrowth in cultured dorsal root ganglia (DRG) cells was significantly inhibited by AT2 receptor antagonist (PD123,319) treatment. AT2 receptor knockdown also inhibited NGF-mediated neurite outgrowth. To determine the mechanisms, we analyzed the NO-cGMP pathway. The cGMP analog increased NGF-mediated nerve elongation, which inhibited by PD123,319. Furthermore, soluble guanylate cyclase expression was significantly less in NGF and PD123,319 treatment DRG than in NGF treatment alone. These results suggest that NGF-mediated neurite outgrowth is suppressed by AT2 receptor signaling via the NO-cGMP-PKG pathway.

αCGRP Transgenic Mice Display Typical Physiologic Features

 Calcitonin gene-related peptide (CGRP) plays an important role in several physiological processes such as vasodilation, cardiovascular homeostasis and transmission of pain. Here we report the generation of a transgenic mouse overexpressing αCGRP and the impact of this on baseline physiological responses. αCGRP transgenic mice displayed significantly increased αCGRP mRNA levels in the kidney, heart and hippocampus. To assess cardiovascular physiology, we measured arterial pressure using a tail cuff system. Heart rate, systolic pressure, mean arterial pressure and diastolic pressure were significantly lower in αCGRP transgenic mice than wild-type mice. To assess pain, a hot plate test was performed and the latency of response was used as an indicator of supraspinal response. In addition, a tail immersion test was performed to assess thermal nociception. A significant increase in latency was observed in the αCGRP transgenic mice when compared with wild-type mice in both tests. These results suggest that αCGRP overexpression causes an increase in thermal reaction and downregulation of the cardiovascular system, presumably due in increased levels of αCGRP.

Effects of alcoholic beverage treatment on spatial learning and fear memory in mice

ABSTRACT Although chronic ethanol treatment is known to impair learning and memory, humans commonly consume a range of alcoholic beverages. However, the specific effects of some alcoholic beverages on behavioral performance are largely unknown. The present study compared the effects of a range of alcoholic beverages (plain ethanol solution, red wine, sake and whiskey; with a matched alcohol concentration of 10%) on learning and memory. 6-week-old C57BL6J mice were orally administered alcohol for 7 weeks. The results revealed that red wine treatment exhibited a trend toward improvement of spatial memory and advanced extinction of fear memory. Additionally, red wine treatment significantly increased mRNA levels of brain-derived neurotrophic factor (BDNF) and N-methyl-D-aspartate (NMDA) receptors in mice hippocampus. These results support previous reports that red wine has beneficial effects. 7-week treatment of red-wine exhibited a trend toward improvement of spatial memory and advanced fear memory extinction with increased BDNF and NMDA receptor mRNA.

The relationship between calcitonin gene-related peptide and depression-like behavior.

痛 み と 情 動 の 新 た な 展 開 3 要約:カルシトニン遺伝子関連ペプチド(CGRP)は, 様々な生理機能を有するペプチドである.強力な血管 弛緩作用がその機能として第一にあげられる.CGRP は神経細胞で産生されて遊離されることから,神経系 との関わりが考えられる.今回は行動異常とCGRPと の関連性について紹介を行う.我々はマウスに 15 日 間ストレスを負荷することによって,海馬における Cgrp mRNA量の減少が起こっていることを明らかに した.そこで,CGRPがうつ様行動に影響を及ぼすの ではないかと仮説を立て,ストレスを負荷するマウス に CGRP(0.5 nmol)を脳室内投与した結果,うつ様 行動の抑制が見られた.同様に海馬新生細胞数の減少 の抑制,神経成長因子(NGF)mRNA量の増加が見ら れた.In vitroにおいても,マウス海馬 E14 培養細胞 株に CGRP(100 nM)を添加すると,Ngf mRNAレベ ルの増加が見られた.そこで,ストレス負荷マウスに NGF受容体阻害薬であるK252aを投与し,行動を観察 した結果,CGRPによる抗うつ作用の減少が観察され た.これらの結果は CGRPが NGFの増加を介して抗 うつ作用を引き起こしている可能性を示唆している.