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三井 烈ミツイ レツ

所属部署医学研究科細胞生理学分野
職名講師
メールアドレス
ホームページURLhttp://scholar.google.com/citations?user=XJ0_pQwAAAAJ&hl=en
生年月日
Last Updated :2020/06/02

研究者基本情報

学歴

  •  - 2000年, 早稲田大学, 人間科学部

学位

  • 静岡県立大学/博士(環境科学)

所属学協会

  • 日本平滑筋学会
  • 日本生理学会
  • 日本解剖学会

経歴

  • 2011-2016 名古屋市立大学大学院医学研究科 助教
  • 2008-2011 早稲田大学大学院人間科学研究科 助教
  • 2006-2008 日本大学松戸歯学部 助手(専任扱)

研究活動情報

研究分野

  • ライフサイエンス, 生理学

研究キーワード

    血管平滑筋, 平滑筋, 自律神経系

論文

  • Synchrony of spontaneous Ca2+ activity in microvascular mural cells., Mitsui R, Hashitani H, Journal of Smooth Muscle Research,   2020年, 査読有り
  • Neural regulation of the contractility of nutrient artery in the guinea-pig tibia., Fukuta H, Mitsui R, Takano H, Hashitani H, Pflügers Archiv-European Journal of Physiology,   2020年, 査読有り
  • Contractile elements and their sympathetic regulations in the pig urinary bladder: a species and regional comparative study., Mitsui R, Lee K, Uchiyama A, Hayakawa S, Kinoshita F, Kajioka S, Eto M, Hashitani H, Cell & Tissue Research, 379, 373 - 387,   2020年, 査読有り
  • Role of K+ channels in maintaining the synchrony of spontaneous Ca2+ transients in the mural cells of rat rectal submucosal arterioles., Mitsui R, Hashitani H, Pflügers Archiv-European Journal of Physiology, 471, 1025 - 1040,   2019年, 査読有り
  • Exercise-induced sympathetic dilatation in arterioles of the guinea pig tibial periosteum., Fukuta H, Mitsui R, Takano H, Hashitani H, Autonomic Neuroscience, 217, 7 - 17,   2019年, 査読有り
  • Role of capillary pericytes in the integration of spontaneous Ca2+ transients in the suburothelial microvasculature in situ of the mouse bladder., Hashitani H, Mitsui R, Miwa-Nishimura K, Lam M, The Journal of Physiology (London), 596, 3531 - 3552,   2018年, 査読有り
  • Mechanisms of PTHrP-induced inhibition of smooth muscle contractility in the guinea pig gastric antrum, H. Ohguchi, R. Mitsui, K. Imaeda, T. Joh, H. Hashitani, NEUROGASTROENTEROLOGY AND MOTILITY, 29, (12) ,   2017年12月, BackgroundParathyroid hormone-related protein (PTHrP) that causes hypercalcemia of malignancy appears to function as an endogenous smooth muscle relaxant. For example, PTHrP released upon bladder wall distension relaxes detrusor smooth muscle to accommodate urine. Here, we explored mechanisms underlying PTHrP-induced suppression of the smooth muscle contractility in the gastric antrum that also undergoes a passive distension. MethodsEffects of PTHrP on phasic contractions and electrical slow waves in the antral smooth muscle of the guinea pig stomach were studied using isometric tension and intracellular microelectrode recordings, respectively. Fluorescent immunohistochemistry was also carried out to identify the distribution of PTH/PTHrP receptors. Key ResultsParathyroid hormone-related protein (1-100nM) reduced the amplitude of phasic contractions and the basal tension. N-nitro-l-arginine (L-NA, 100M), a nitric oxide (NO) synthase inhibitor, or 1H-[1,2,4]oxadiazolo-[4, 3-a]quinoxalin-1-one (ODQ, 10 mu M), a guanylate cyclase inhibitor, diminished the PTHrP (10nM)-induced reduction in the amplitude of phasic contractions. SQ22536 (300M), an adenylate cyclase inhibitor, attenuated the PTHrP-induced reduction in basal tension. The combination of ODQ (10M) and SQ22536 (300M) inhibited the PTHrP-induced reductions in both phasic contractions and basal tension. PTHrP (100nM) had no inhibitory effect on the electrical slow waves in the antral smooth muscle. PTH/PTHrP receptors were expressed in cell bodies of PGP9.5-positive neurons in the myenteric plexus. Conclusions & InferencesParathyroid hormone-related protein exerts its inhibitory actions on the antral smooth muscle via both nitric oxide-cyclic guanosine monophosphate (NO-cGMP) and cyclic adenosine monophosphate (AMP) pathways. Thus, PTHrP may act as an endogenous relaxant of the gastric antrum employing the two complementary signaling pathways to ensure the adaptive relaxation of stomach.
  • Nerve-induced responses of mouse vaginal smooth muscle, Dirk F. van Helden, Ayumi Kamiya, Sam Kelsey, Derek R. Laver, Phillip Jobling, Retsu Mitsui, Hikaru Hashitani, PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 469, (10) 1373 - 1385,   2017年10月, Neural and agonist-induced contractions of proximal (i.e. upper half adjacent to the cervix) and distal mouse vaginal smooth muscle strips were investigated. We hypothesised that nerve-mediated vaginal contractions arise through activity of cholinergic nerves. Nerve activation by bursts of electrical field stimulation (EFS) caused a primary transient contraction often accompanied by a secondary transient contraction, both larger in proximal than distal tissues (i.e. primary: 7-fold larger; secondary: 3-fold larger). Our hypothesis was supported as we found that cholinergic nerves mediated the primary transient contraction in both proximal and distal vaginal strips, as EFS responses were enhanced by neostigmine an anticholinesterase, massively inhibited by the competitive muscarinic receptor antagonist atropine and not affected by the non-selective alpha-adrenergic receptor antagonist phentolamine. Primary transient contractions were halved in amplitude by the L-type Ca2+ channel blocker nifedipine and markedly inhibited by the sarco-endoplasmic reticulum calcium ATPase (SERCA) inhibitor cyclopiazonic acid (CPA). Resultant secondary transient contractions were abolished by nifedipine. Notably, the selective alpha(1)-adrenergic receptor agonist phenylephrine caused tonic contracture in distal but not proximal strips. Low-frequency EFS often initiated recurrent transient contractions similar to those elicited by CCh. Immunohistochemical studies demonstrated innervation of the smooth muscle. Findings of enhanced proximal cholinergic nerve-induced transient contractions, evidence that maintained nerve stimulation could cause recurrent contractions and the finding of distal phenylephrine-mediated tonic contraction have implications on insemination.
  • Contractile properties of periosteal arterioles in the guinea-pig tibia, Hiroyasu Fukuta, Retsu Mitsui, Hiromichi Takano, Hikaru Hashitani, PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 469, (9) 1203 - 1213,   2017年09月, The periosteal arterioles of the compact bone may play a critical role in bone growth. To explore the contractile properties of tibial arterioles, spontaneous and nerve-evoked constrictions were compared in preparations from 3-week-old and 1-year-old guinea-pigs. Changes in arteriole diameters were measured using video microscopy. Their innervation was investigated using fluorescence immunohistochemistry. Fifty per cent and 40% of tibial arterioles from 3-week-old and 1-year-old guinea-pigs, respectively, exhibited spontaneous phasic constrictions that were inhibited by 1 mu M nifedipine, 10 mu M cyclopiazonic acid or 100 mu M 2-APB. Nerve-evoked phasic constrictions in both age groups were largely suppressed by phentolamine (1 mu M), an alpha-adrenoceptor antagonist, or sympathetic neurotransmitter depletion using guanethidine (10 mu M) but were enhanced by spanttide (1 mu M), a substance P receptor antagonist, or L-nitro arginine (L-NA; 100 mu M), an inhibitor of nitric oxide synthase (NOS). Nerve-evoked constrictions in 1-year-old animals were smaller than those in younger animals but greatly enhanced by L-NA. Immunohistochemistry revealed sympathetic and substance P-positive primary afferent nerves running along the arterioles as well as endothelial NOS expression in both age groups. Spontaneous arteriolar constrictions appear to rely on both Ca2+ release from the sarcoplasmic reticulum and Ca2+ influx through L-type Ca2+ channels. Noradrenaline released from sympathetic nerves triggers arteriolar constriction, while substance P released from primary afferent nerves dilates the arterioles by releasing nitric oxide (NO), presumably from the endothelium. Thus, the enhanced endothelial NO release in adult guinea-pigs may be important to increase the blood supply to meet the increased metabolic demands during bone growth.
  • Properties of synchronous spontaneous Ca2+ transients in the mural cells of rat rectal arterioles, Retsu Mitsui, Hikaru Hashitani, PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 469, (9) 1189 - 1202,   2017年09月, Synchrony of spontaneous Ca2+ transients among venular mural cells (smooth muscle cells and pericytes) in visceral organs relies on the intercellular spread of L-type voltage-dependent Ca2+ channel (LVDCC)-dependent depolarisations. However, the mechanisms underlying the synchrony of spontaneous Ca2+ transients between arteriolar mural cells are less understood. The spontaneous intracellular Ca2+ dynamics of arteriolar mural cells in the rat rectal submucosa were visualised by Cal-520 Ca2+ imaging to analyse their synchrony. The mural cells in fine arterioles that had a rounded cell body with several extended processes developed spontaneous 'synchronous' Ca2+ transients arising from Ca2+ released from sarcoendoplasmic reticulum Ca2+ stores. Gap junction blockers (3 mu M carbenoxolone, 10 mu M 18 beta-glycyrrhetinic acid), a Ca2+-activated Cl- channel (CaCC) blocker (100 mu M 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid) or lowering extracellular Cl- concentration (from 134.4 to 12.4 mM) disrupted the synchrony of Ca2+ transients between arteriolar mural cells. Blockers of T-type voltage-dependent Ca2+ channels (TVDCCs, 1 mu M mibefradil or ML218) or LVDCCs (1 mu M nifedipine) reduced the Ca2+ transient frequency or their area under curve (AUC), respectively. However, neither TVDCC nor LVDCC blockers disrupted the synchrony of Ca2+ transients among arteriolar mural cells. This is in contrast with rectal venules in which nifedipine disrupted the synchrony of spontaneous Ca2+ transients. Thus, spontaneous transient depolarisations arising from the opening of CaCCs may effectively spread to neighbouring arteriolar mural cells via gap junctions to maintain the Ca2+ transient synchrony. Activation of TVDCCs appears to accelerate spontaneous Ca2+ transients, while LVDCCs predominantly contribute to the duration of Ca2+ transients.
  • Interstitial cell modulation of pyeloureteric peristalsis in the mouse renal pelvis examined using FIBSEM tomography and calcium indicators, Hikaru Hashitani, Michael J. Nguyen, Haruka Noda, Retsu Mitsui, Ryuhei Higashi, Keisuke Ohta, Kei-Ichiro Nakamura, Richard J. Lang, PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 469, (5-6) 797 - 813,   2017年06月, Typical and atypical smooth muscle cells (TSMCs and ASMCs, respectively) and interstitial cells (ICs) within the pacemaker region of the mouse renal pelvis were examined using focused ion beam scanning electron (FIB SEM) tomography, immunohistochemistry and Ca2+ imaging. Individual cells within 500-900 electron micrograph stacks were volume rendered and associations with their neighbours established. 'Ribbon-shaped', Ano1 Cl- channel immuno-reactive ICs were present in the adventitia and the sub-urothelial space adjacent to the TSMC layer. ICs in the proximal renal pelvis were immuno-reactive to antibodies for Ca(V)3.1 and hyperpolarization-activated cation nucleotide-gated isoform 3 (HCN3) channel sub-units, while basal-epithelial cells (BECs) were intensely immuno-reactive to Kv7.5 channel antibodies. Adventitial to the TSMC layer, ASMCs formed close appositions with TSMCs and ICs. The T-type Ca(2+)channel blocker, Ni2+ (10-200 mu M), reduced the frequency while the L-type Ca2+ channel blocker (1 mu M nifedipine) reduced the amplitude of propagating Ca2+ waves and contractions in the TSMC layer. Upon complete suppression of Ca2+ entry through TSMC Ca2+ channels, ASMCs displayed high-frequency (6 min(-1)) Ca2+ transients, and ICs distributed into two populations of cells firing at 1 and 3 min(-1), respectively. IC Ca2+ transients periodically (every 3-5 min(-1)) summed into bursts which doubled the frequency of ASMC Ca2+ transient firing. Synchronized IC bursting and the acceleration of ASMC firing were inhibited upon blockade of HCN channels with ZD7288 or cell-to-cell coupling with carbenoxolone. While ASMCs appear to be the primary pacemaker driving pyeloureteric peristalsis, it was concluded that sub-urothelial HCN3(+), Ca(V)3.1(+) ICs can accelerate ASMC Ca2+ signalling.
  • Role of PTHrP and Sensory Nerve Peptides in Regulating Contractility of Muscularis Mucosae and Detrusor Smooth Muscle in the Guinea Pig Bladder, Ken Lee, Retsu Mitsui, Shunichi Kajioka, Seiji Naito, Hikaru Hashitani, JOURNAL OF UROLOGY, 196, (4) 1287 - 1294,   2016年10月, Purpose: We investigated the neurohumoral modulation of the contractility of bladder muscularis mucosae (mucosa) compared with that of detrusor smooth muscle. Materials and Methods: Changes in the contractility of mucosal and detrusor bundles from guinea pig bladders were measured using isometric tension recording. The morphological relationship between the muscularis mucosae and blood vessels, and their sensory innervation was examined by fluorescence immunohistochemistry. Results: Meshworks of muscularis mucosae with numerous branches and anastomosis preferentially ran parallel with suburothelial blood vessels. Although PTHrPRs (parathyroid hormone-related peptide receptors) were expressed in detrusor and mucosa, the endogenous detrusor relaxant PTHrP (parathyroid hormone-related peptide) (1 nM) suppressed spontaneous contractions in detrusor but not in mucosa. A higher concentration of PTHrP (10 nM) was required to inhibit mucosal contractility. Capsaicin (1 mu M) abolished spontaneous contractions in mucosa but had an excitatory action on detrusor contractility. hCGRP (human calcitonin gene-related peptide) (1 nM) attenuated spontaneous mucosal contractions. Pretreatment with the CGRP (calcitonin gene-related peptide) antagonist hCGRP 8-37 (2 mu M) inhibited CGRP or capsaicin induced suppression of spontaneous contractions. Consistently, CGRP immunoreactive primary afferent nerves were abundant in muscularis mucosae. Conclusions: Co-localization of muscularis mucosae with the suburothelial microvasculature suggests that spontaneous contractions of mucosa might function to prevent microvasculature stretching upon bladder wall distension during the storage phase. It is likely that PTHrP selectively suppresses spontaneous contractions in detrusor but not in mucosa. Thus, endogenous PTHrP may well increase bladder compliance without an associated distension induced deformation of mucosal elements. Excessive stimulations of sensory nerves may suppress mucosal contractility by releasing CGRP.
  • Functional coupling of TRPV4 channels and BK channels in regulating spontaneous contractions of the guinea pig urinary bladder, Ayu Isogai, Ken Lee, Retsu Mitsui, Hikaru Hashitani, PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 468, (9) 1573 - 1585,   2016年09月, We investigated the role of TRPV4 channels (TRPV4) in regulating the contractility of detrusor smooth muscle (DSM) and muscularis mucosae (MM) of the urinary bladder. Distribution of TRPV4 in DSM and MM of guineapig bladders was examined by fluorescence immunohistochemistry. Changes in the contractility of DSM and MM bundles were measured using isometric tension recording. Intracellular Ca2+ dynamics were visualized by Cal-520 fluorescent Ca2+ imaging, while membrane potential changes were recorded using intracellular microelectrode technique. DSM and MM expressed TRPV4 immunoreactivity. GSK1016790A (GSK, 1 nM), a TRPV4 agonist, evoked a sustained contraction in both DSM and MM associated with a cessation of spontaneous phasic contractions in a manner sensitive to HC-067047 (10 mu M), a TRPV4 antagonist. Iberiotoxin (100 nM) and paxilline (1 mu M), large conductance Ca2+-activated K+ (BK) channel blockers restored the spontaneous contractions in GSK. The sustained contractions in DSMandMMwere reduced by nifedipine (10 mu M), a blocker of L-type voltage-dependent Ca2+ channels (LVDCCs) by about 40 % and by nominally Ca2+-free solution by some 90 %. GSK (1 nM) abolished spontaneous Ca2+ transients, increased basal Ca2+ levels and also prevented spontaneous action potential discharge associated with DSM membrane hyperpolarization. In conclusion, Ca2+ influx through TRPV4 appears to activate BK channels to suppress spontaneous contractions and thus a functional coupling of TRPV4 with BK channels may act as a self-limiting mechanism for bladder contractility during its storage phase. Despite the membrane hyperpolarization in GSK, Ca2+ entry mainly through TRPV4 develops the tonic contraction.
  • Mechanisms underlying spontaneous constrictions of postcapillary venules in the rat stomach, Retsu Mitsui, Hikaru Hashitani, PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 468, (2) 279 - 291,   2016年02月, Postcapillary venules (PCVs) play a critical role in regulating capillary hydrostatic pressure, but their contractile mechanisms are not well understood. We examined the properties of spontaneous vasomotion and corresponding Ca2+ transients in gastric PCV. In the rat gastric submucosa, changes in PCV diameter and intracellular Ca2+ dynamics were visualised by video tracking system and fluorescent Ca2+ imaging, respectively, while PCV morphology was examined by immunohistochemistry. Stellate-shaped PCV mural cells expressing alpha-smooth muscle actin exhibited synchronised spontaneous Ca2+ transients to develop vasomotion which was abolished by nifedipine (1 mu M), cyclopiazonic acid (10 mu M), or Ca2+-activated Cl- channel inhibitors (100 mu M niflumic acid, 1 mu M T16Ainh-A01). A gap junction blocker (3 mu M carbenoxolone) disrupted the synchrony of spontaneous Ca2+ transients amongst PCV mural cells and attenuated spontaneous vasomotion. Low chloride solution ([Cl-](0) = 12.4 mM) also disrupted the synchrony of spontaneous Ca2+ transients and abolished vasomotion. Na+-K+-Cl- co-transporter inhibitors (10 mu M bumetanide, 30 mu M furosemide) suppressed spontaneous Ca2+ transients and vasoconstrictions. A phosphodiesterase type 5 (PDE5) inhibitor (1 mu M tadalafil) disrupted the spontaneous Ca2+ transient synchrony and abolished vasomotion in a nitric oxide (NO)-dependent manner. Thus, gastric PCVs exhibit spontaneous vasomotion, resulting from synchronised spontaneous Ca2+ transients within a network of stellate-shaped PCV mural cells. An active Cl- accumulation partly via Na+-K+-Cl- co-transport appears to be fundamental in maintaining depolarisation upon the opening of Ca2+-activated Cl- channels that triggers Ca2+ influx via voltage-dependent L-type Ca2+ channels. Basal PDE5 activity may continuously counteract vaso-relaxing effects of endothelial NO to maintain spontaneous vasomotion.
  • Electrical properties of purinergic transmission in smooth muscle of the guinea-pig prostate, Michelle Lam, Retsu Mitsui, Hikaru Hashitani, AUTONOMIC NEUROSCIENCE-BASIC & CLINICAL, 194, 8 - 16,   2016年01月, Prostatic smooth muscle develops spontaneous myogenic tone which is modulated by autonomic neuromuscular transmission. This study aimed to investigate the role of purinergic transmission in regulating electrical activity of prostate smooth muscle and whether its contribution may be altered with age. Intracellular recordings were simultaneously made with isometric tension recordings in smooth muscle preparations of the guinea-pig prostate. Immunostaining for P2X1 receptors on whole mount preparations was also performed. In prostate preparations which generated spontaneous slow waves, electrical field stimulation (EFS)-evoked excitatory junction potentials (EJPs) which were abolished by guanethidine (10 mu M), alpha-beta-methylene ATP (10 mu M) or pyridoxal phosphate-6-azophenyl-2,4-disulfonic acid (PPADS, 10 mu M) but not phentolamine (1 mu M). Consistently, immunostaining revealed the expression of P2X1 receptors on prostatic smooth muscle. EJPs themselves did not cause contractions, but EJPs could sum to trigger a slow wave and associated contraction. Yohimbine (1 mu M) and 3,7-dimethyl-1-propargylxanthine (DMPX, 10 mu M) but not propranolol (1 mu M) potentiated EJPs. Although properties of EJPs were not different between young and aging guinea-pig prostates, ectoATPase inhibitor ARL 67156 (100 mu M) augmented EJP amplitudes by 64.2 +/- 29.6% in aging animals, compared to 22.1 +/- 19.9% in young animals. These results suggest that ATP released from sympathetic nerves acts on P2X1 purinoceptors located on prostate smooth muscle to evoke EJPs, while pre-junctional alpha(2)-adrenergic and adenosine A(2) receptors may play a role in preventing excessive transmitter release. Age-related up-regulation of enzymatic ATP breakdown may be a compensatory mechanism for the enhanced purinergic transmission which would cause hypercontractility arising from increased ATP release in older animals. (C) 2015 Elsevier B.V. All rights reserved.
  • Pacemaker role of pericytes in generating synchronised spontaneous Ca2+ transients in the myenteric microvasculature of the guinea-pig gastric antrum., Hashitani H, Mitsui R, Masaki S, van Helden DF, Cell Calcium, 58, 442 - 456,   2015年
  • Functional properties of submucosal venules in the rat stomach., Mitsui R, Hashitani H, Pflügers Archiv - European Journal of Physiology, 467, 1327 - 1342,   2015年
  • Characterization of slow waves generated by myenteric interstitial cells of Cajal of the rabbit small intestine., Kito Y, Mitsui R, Ward SM, Sanders KM, American Journal of Physiology-Gastrointestinal and Liver Physiology, 308, G378 - G388,   2015年
  • Spontaneous transient hyperpolarizations in the rabbit small intestine, Yoshihiko Kito, Masaaki Kurahashi, Retsu Mitsui, Sean M. Ward, Kenton M. Sanders, JOURNAL OF PHYSIOLOGY-LONDON, 592, (21) 4733 - 4745,   2014年11月, Recently, it was shown that fibroblast-like cells (FLCs) possess the apparatus to mediate purinergic motor neurotransmission in the gastrointestinal tract. However, the electrophysiological properties of FLCs in situ have not been determined. We recorded two patterns of slow waves from longitudinal smooth muscle cells and circular smooth muscle cells, large amplitude slow waves from interstitial cells of Cajal, and spontaneous transient hyperpolarizations (STHs) from FLCs in the rabbit small intestine using intracellular recording combined with dye injection to identify the cellular morphology of impaled cells. Drugs that inhibit the signalling pathway involved in purinergic neurotransmission inhibited STHs in FLCs. Small amplitude STHs were recorded in smooth muscle cells but not in interstitial cells of Cajal, suggesting that STHs from FLCs were conducted passively to smooth muscle cells. We conclude that FLCs display the molecular apparatus necessary to mediate purinergic neurotransmission and may tonically dampen smooth muscle excitability in the rabbit small intestine by an ongoing discharge of STHs. AbstractFour types of electrical activity were recorded and related to cell structure by intracellular recording and dye injection into impaled cells in muscles of rabbit small intestine. The specific cell types from which recordings were made were longitudinal smooth muscle cells (LSMCs), circular smooth muscle cells (CSMCs), interstitial cells of Cajal distributed in the myenteric region (ICC-MY) and fibroblast-like cells (FLCs). Slow waves (slow waves(SMC)) were recorded from LSMCs and CSMCs. Slow waves (slow waves(ICC)) were of greatest amplitude (>50mV) and highest maximum rate of rise (>10Vs(-1)) in ICC-MY. The dominant activity in FLCs was spontaneous transient hyperpolarizations (STHs), with maximum amplitudes above 30mV. STHs were often superimposed upon small amplitude slow waves (slow waves(FLC)). STHs displayed a cyclical pattern of discharge irrespective of background slow wave activity. STHs were inhibited by MRS2500 (3m), a P2Y1 antagonist, and abolished by apamin (0.3m), a blocker of small conductance Ca2+-activated K+ channels. Small amplitude STHs (<15mV) were detected in smooth muscle layers, whereas STHs were not resolved in cells identified as ICC-MY. Electrical field stimulation evoked purinergic inhibitory junction potentials (IJPs) in CSMCs. Purinergic IJPs were not recorded from ICC-MY. These results suggest that FLCs may regulate smooth muscle excitability in the rabbit small intestine via generation of rhythmic apamin-sensitive STHs. Stimulation of P2Y1 receptors modulates the amplitudes of STHs. Our results also suggest that purinergic inhibitory motor neurons regulate the motility of the rabbit small intestine by causing IJPs in FLCs that conduct to CSMCs.
  • Voltage Dependence of Slow Wave Frequency in the Guinea Pig Prostate, Yusuke Shigemasa, Michelle Lam, Retsu Mitsui, Hikaru Hashitani, JOURNAL OF UROLOGY, 192, (4) 1286 - 1292,   2014年10月, Purpose: Spontaneous phasic contractions of the guinea pig prostate stroma result from the generation of slow waves that appear to primarily rely on spontaneous Ca2+ release from the endoplasmic/sarcoplasmic reticulum and subsequent opening of Ca2+ activated chloride channels. We investigated voltage dependent mechanisms in the regulation of slow wave frequency. Materials and Methods: Changes in membrane potential were recorded using conventional intracellular recording techniques while simultaneously measuring the isometric tension of guinea pig prostate lobes. Fluorescence immunohistochemistry was done to determine the cellular composition of the prostate stroma. Results: Depolarization induced by high K+ solution, K+ free solution or outward current injection was associated with increased slow wave frequency. In contrast, hyperpolarization induced by the re-addition of K+, adenosine triphosphate sensitive K+ channel openers or inward current injection prevented slow wave generation. K+ channel openers induced hyperpolarization and the cessation of slow waves was reversed by glibenclamide (10 mu M). Nifedipine (1 to 10 mu M) shortened the duration of slow waves and pacemaker potentials but often failed to prevent their generation and associated contractions. Subsequently Ni2+ (100 mu M) or mibefradil (1 mu M) largely suppressed slow waves and abolished residual contractions. Immunohistochemistry revealed small interconnected smooth muscle bundles as well as vimentin positive interstitial cells but failed to show a network of Kit positive interstitial cells. Conclusions: Prostate slow wave frequency is voltage dependent due to the significant contribution of L-type and T-type Ca2+ channels. Prostate slow waves may arise from cooperation between spontaneous Ca2+ release from internal stores and plasmalemmal voltage dependent Ca2+ channels.
  • Neurohumoral regulation of spontaneous constrictions in suburothelial venules of the rat urinary bladder, Yuki Shimizu, Satoshi Mochizuki, Retsu Mitsui, Hikaru Hashitani, VASCULAR PHARMACOLOGY, 60, (2) 84 - 94,   2014年02月, Venules of the bladder suburothelium develop spontaneous phasic constrictions that may play a critical role in maintaining venular drainage of tissue metabolites. We aimed to investigate neurohumoral regulation of the spontaneous venular constrictions (SVCs). Changes in venular diameter of the rat bladder suburothelium were monitored using a video tracking system, whilst the effects of electrical field stimulation (EFS) and bathapplied bioactive substances were investigated. The innervation of the suburothelial microvasculature was examined by immunohistochemistry. EFS (10 Hz for 30 s) induced an increase in the frequency of SVCs that was prevented by phentolamine (1 mu M). In phentolamine-pretreated venules, EFS suppressed SVCs with a venular dilatation in a manner attenuated by propranolol (1 mu M) or L-nitro arginine (LNA, 10 mu M). BRL37344 (1 mu M), a beta(3) adrenoceptor agonist, dilated venules and reduced the frequency of SVCs in an LNA-sensitive manner. ACh (1-10 mu M) increased the frequency of SVCs. ATP (1 mu M) transiently constricted venules and then caused LNA-sensitive cessation of SVCs associated with a dilatation. Substance P (100 nM) caused a venular constriction, whilst calcitonin gene related peptide (CGRP, 100 nM) caused a dilatation of venules and suppression of SVCs that were not inhibited by LNA. Immunohistochemical staining demonstrated sympathetic as well as substance P- and CGRP-containing nerves running along the venules. Spontaneous constrictions of suburothelial venules are accelerated by sympathetic alpha-adrenergic stimulation, but suppressed upon beta-adrenergic stimulation. In addition, suburothelial venular constrictions appear to be modulated by several bioactive substances that could be released from urothelium or suburothelial sensory nerves. (C) 2014 Elsevier Inc. All rights reserved.
  • Immunohistochemical characteristics of suburothelial microvasculature in the mouse bladder, Retsu Mitsui, Hikaru Hashitani, HISTOCHEMISTRY AND CELL BIOLOGY, 140, (2) 189 - 200,   2013年08月, The morphological characteristics of smooth muscle cells (SMCs) and their innervation of the suburothelial microvasculature of the mouse bladder were investigated by immunohistochemistry. Whole mount bladder mucosal preparations were immune-stained for alpha-smooth muscle actin (alpha-SMA) and/or neuronal markers and examined using confocal laser scanning microscopy. Suburothelial arterioles consisted of alpha-SMA-immunopositive circular smooth muscle cells, while the venular wall composed of alpha-SMA-positive SMCs that displayed several processes which extended from their cell bodies to form an extensive meshwork. In larger venules, a complex meshwork of stellate-shaped SMCs were observed. NG2 chondroitin sulphate proteoglycan-immunoreactive cell bodies of capillary pericytes were not immunoreactive for alpha-SMA. In the rat bladder suburothelial venules, circular SMCs were the dominant cell type expressing alpha-SMA-immunoreactivity. Since alpha-SMA-positive SMCs in suburothelial arterioles and venules in the mouse bladder had quite distinct morphologies, the innervation of both vessels could be examined by double labelling for alpha-SMA and various neuronal markers. Varicose nerve bundles immunoreactive for tyrosine hydroxylase (sympathetic nerves), choline acetyltransferase (cholinergic nerves) or substance P (primary afferent nerves) were all detected along side suburothelial arterioles. Single varicose nerve fibres positive for these three neuronal markers were also detected around the venules. Thus, whole mount preparations are useful when examining the morphology of alpha-SMA-positive SMCs of the microvasculature in the suburothelium of mouse bladder as well as their relationship with their innervations. In conclusion, arterioles and venules of the bladder suburothelium are the target of sympathetic, cholinergic and primary afferent nerve fibres.
  • Properties of submucosal venules in the rat distal colon., Mitsui R, Miyamoto S, Takano H, Hashitani H, British Journal of Pharmacology, 170, 968 - 977,   2013年
  • Functional and morphological properties of pericytes in suburothelial venules of the mouse bladder, Hikaru Hashitani, Retsu Mitsui, Yuki Shimizu, Ryuhei Higashi, Keiichiro Nakamura, BRITISH JOURNAL OF PHARMACOLOGY, 167, (8) 1723 - 1736,   2012年12月, Background and Purpose In suburothelial venules of rat bladder, pericytes (perivascular cells) develop spontaneous Ca2+ transients, which may drive the smooth muscle wall to generate spontaneous venular constrictions. We aimed to further explore the morphological and functional characteristics of pericytes in the mouse bladder. Experimental Approach The morphological features of pericytes were investigated by electron microscopy and fluorescence immunohistochemistry. Changes in diameters of suburothelial venules were measured using video microscopy, while intracellular Ca2+ dynamics were visualized using Fluo-4 fluorescence Ca2+ imaging. Key Results A network of a-smooth muscle actin immunoreactive pericytes surrounded venules in the mouse bladder suburothelium. Scanning electron microscopy revealed that this network of stellate-shaped pericytes covered the venules, while transmission electron microscopy demonstrated that the venular wall consisted of endothelium and adjacent pericytes, lacking an intermediate smooth muscle layer. Pericytes exhibited spontaneous Ca2+ transients, which were accompanied by phasic venular constrictions. Nicardipine (1?mu M) disrupted the synchrony of spontaneous Ca2+ transients in pericytes and reduced their associated constrictions. Residual asynchronous Ca2+ transients were suppressed by cyclopiazonic acid (10?mu M), 2-aminoethoxydiphenyl borate (10?mu M), U-73122 (1?mu M), oligomycin (1?mu M) and SKF96365 (10?mu M), but unaffected by ryanodine (100?mu M) or YM-244769 (1?mu M), suggesting that pericyte Ca2+ transients rely on Ca2+ release from the endoplasmic reticulum via the InsP3 receptor and also require Ca2+ influx through store-operated Ca2+ channels. Conclusions and Implications The pericytes in mouse bladder can generate spontaneous Ca2+ transients and contractions, and thus have a fundamental role in promoting spontaneous constrictions of suburothelial venules.
  • Potassium and ANO1/TMEM16A chloride channel profiles distinguish atypical and typical smooth muscle cells from interstitial cells in the mouse renal pelvis, Javed Iqbal, Mary A. Tonta, Retsu Mitsui, Qun Li, Michelle Kett, Jinhua Li, Helena C. Parkington, Hikaru Hashitani, Richard J. Lang, BRITISH JOURNAL OF PHARMACOLOGY, 165, (7) 2389 - 2408,   2012年04月, BACKGROUND AND PURPOSE Although atypical smooth muscle cells (SMCs) in the proximal renal pelvis are thought to generate the pacemaker signals that drive pyeloureteric peristalsis, their location and electrical properties remain obscure. EXPERIMENTAL APPROACH Standard patch clamp, intracellular microelectrode and immunohistochemistry techniques were used. To unequivocally identify SMCs, transgenic mice with enhanced yellow fluorescent protein (eYFP) expressed in cells containing alpha-smooth muscle actin (alpha-SMA) were sometimes used. KEY RESULTS Atypical SMCs were distinguished from typical SMCs by the absence of both a transient 4-aminopyridine-sensitive K+ current (I-KA) and spontaneous transient outward currents (STOCs) upon the opening of large-conductance Ca2+-activated K+ (BK) channels. Many typical SMCs displayed a slowly activating, slowly decaying Cl- current blocked by niflumic acid (NFA). Immunostaining for K(V)4.3 and ANO1/TMEM16A Cl- channel subunits co-localized with alpha-SMA immunoreactive product predominately in the distal renal pelvis. Atypical SMCs fired spontaneous inward currents that were either selective for Cl- and blocked by NFA, or cation-selective and blocked by La3+. alpha-SMA(-) interstitial cells (ICs) were distinguished by the presence of a Xe991-sensitive K(V)7 current, BK channel STOCs and Cl- selective, NFA-sensitive spontaneous transient inward currents (STICs). Intense ANO1/TMEM16A and K(V)7.5 immunostaining was present in Kit(-) alpha-SMA(-) ICs in the suburothelial and adventitial regions of the renal pelvis. CONCLUSIONS AND IMPLICATIONS We conclude that K(V)4.3(+) alpha-SMA(+) SMCs are typical SMCs that facilitate muscle wall contraction, that ANO1/TMEM16A and K(V)7.5 immunoreactivity may be selective markers of Kit(-) ICs and that atypical SMCs which discharge spontaneous inward currents are the pelviureteric pacemakers.
  • Functional Properties of Suburothelial Microvessels in the Rat Bladder, Hikaru Hashitani, Hiromich Takano, Kohei Fujita, Retsu Mitsui, Hikaru Suzuki, JOURNAL OF UROLOGY, 185, (6) 2382 - 2391,   2011年06月, Purpose: We investigated the properties of suburothelial microvessels, which have a vital role in maintaining microcirculation to cells involved in bladder afferent signaling. Materials and Methods: Changes in the diameter of rat bladder suburothelial microvessels were measured using video microscopy. Membrane potential changes and intracellular Ca(2+) dynamics of suburothelial venules were examined using intracellular recording techniques and Ca(2+) imaging of fluo-4 fluorescence, respectively. Results: Suburothelial venules showed spontaneous action potential and vasoconstriction activity while suburothelial arterioles were quiescent. Venular vasoconstriction was prevented by cyclopiazonic acid or nicardipine and decreased by 2-aminoethoxydiphenyl borate, niflumic acid or 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid. Venular smooth muscle cells and perivascular interstitial cells showed spontaneous Ca(2+) transients. Nicardipine decreased the amplitude and disrupted the synchronicity of Ca(2+) transients in and between the 2 cell populations. Residual Ca(2+) transients in nicardipine occurred asynchronously and were abolished by cyclopiazonic acid. Suburothelial arterioles constricted in response to transmural nerve stimulation. These nerve induced constrictions were suppressed by prazosin or the selective alpha(1A) blocker RS17053 but not by the alpha(1D) blocker BMY7378. Remaining constrictions were abolished by guanethidine. Conclusions: Spontaneous vasoconstriction of suburothelial venules appears to result upon Ca(2+) release from the sarcoplasmic reticulum upon activation of inositol 1,4,5-trisphosphate receptors. This Ca(2+) opens Ca(2+) activated Cl(-) channels to trigger action potentials and Ca(2+) influx through L-type Ca(2+) channels. Adjacent perivascular interstitial cells may also have a role in generating this spontaneous venular vasoconstriction. In contrast, sympathetic nerve released noradrenaline acts on alpha(1A)-adrenoceptors to constrict suburothelial arterioles.
  • Immunohistochemical analysis of substance P-containing neurons in rat small intestine, Retsu Mitsui, CELL AND TISSUE RESEARCH, 343, (2) 331 - 341,   2011年02月, The neuropeptide substance P (SP) is involved in the regulation of epithelial secretion and motility in the rat small intestine. The morphology, chemical profiles and proportion of SP-containing enteric neurons in this tissue have been examined by immunohistochemical analysis of whole-mount preparations obtained from colchicine-treated rats. In the submucosal plexus of the duodenum, jejunum and ileum, the proportion of SP-positive neurons is 53%, 51% and 49%, respectively. All SP-positive submucosal neurons are positive for neurofilament 200 (NF-200) and calretinin. Immunoreactivity for calcitonin gene-related peptide (CGRP) is detectable in 55% of the SP-positive submucosal neurons. Some SP-positive submucosal neurons have two or more long processes emerging from an oval or round cell body, a characteristic of the Dogiel type II neuron (type II neuron; a putative intrinsic primary afferent neuron). About one-third of the neurons in the myenteric plexus are positive for SP and a majority of them are NF-200/calretinin-positive type II neurons. Immunoreactivity for the SP receptor neurokinin-1 receptor (NK1R) has been detected mainly in the submucosal and myenteric NF-200-positive neurons, which are expected to contain SP. These neurons possibly stimulate each other via SP release. Most of the submucosal and myenteric neurons, including type II neurons, show immunoreactive for the prostaglandin E2 receptor EP3 receptor (EP3R). Thus, SP/NF-200/calretinin/NK1R/EP3R is the common chemical profile of type II neurons in the rat small intestine. The proportion of SP-immunopositive submucosal neurons (49%-53%) is higher in the rat small intestine than in the colon (a parts per thousand currency sign11%) and around 50% are positive for CGRP.
  • Maintenance of paracellular barrier function by insulin-like growth factor-I in submandibular gland cells, Retsu Mitsui, Junko Fujita-Yoshigaki, Takanori Narita, Miwako Matsuki-Fukushima, Keitaro Satoh, Bing Qi, Ming-Yu Guo, Osamu Katsumata-Kato, Hiroshi Sugiya, ARCHIVES OF ORAL BIOLOGY, 55, (12) 963 - 969,   2010年12月, Insulin-like growth factor-I (IGF-I) is expressed in salivary glands. We examined the effects of IGF-I on cell number, the expression and distribution of tight junction proteins and the paracellular barrier function in cells derived from rat submandibular glands. When those cells were cultured in medium containing 10% foetal bovine serum (FBS) or IGF-I, the number of cells was comparable at 10 days. However, in the presence of inhibitor of IGF-I receptors, the number of cells cultured with FBS only was clearly reduced. The tight junction proteins occludin and claudin-3 were similarly detected by Western blotting in cells cultured with IGF-I or FBS. Immunostaining revealed that occludin and another tight junction protein (ZO-1) were similarly localized at intracellular junctions of cells cultured with IGF-I or FBS. The barrier functions were evaluated by transepithelial resistance (TER) and by FITC-dextran permeability. The TER values and FITC-dextran permeability of cells cultured with IGF-I or FBS were comparable. These observations suggest that IGF-I contributes to the maintenance not only of the cell number of salivary gland cells but also of their paracellular barrier function via the expression and distribution of tight junction proteins. (C) 2010 Elsevier Ltd. All rights reserved.
  • Immunohistochemical characteristics of submucosal Dogiel type II neurons in rat colon, Retsu Mitsui, CELL AND TISSUE RESEARCH, 340, (2) 257 - 265,   2010年05月, Secretory and motility reflexes are evoked by physiological stimuli in the isolated rat distal colon, which is therefore expected to contain intrinsic primary afferent (sensory) neurons. Dogiel type II neurons (putative intrinsic primary afferent neurons) exhibit several long processes emerging from large oval or round cell bodies. This study has examined the immunohistochemical characteristics of type II neurons in the submucosal plexus of rat distal colons by using whole-mount preparations. Neuronal cell bodies positive for both substance P (SP) and calretinin have been observed in colchicine-treated rats. Neurofilament 200 immunostaining has confirmed the type II morphology of SP-positive neurons. Moreover, all submucosal type II neurons identified by neurofilament 200 immunoreactivity are positive for calretinin. Calcitonin gene-related peptide (CGRP)-positive neurons in the submucosal plexus are distinct from type II neurons because they are negative for calretinin and have smaller cell bodies than the SP-positive submucosal type II neurons. Most (73%) of the submucosal neurons including type II neurons exhibit immunoreactivity for the neurokinin-1 receptor (NK1R), a receptor for SP, on the surface of cell bodies. Immunoreactivity for the EP3 receptor (EP3R), a receptor for prostaglandin E2, has been detected in 51% of submucosal neurons including type II neurons. Thus, submucosal type II neurons in the rat distal colon are immunopositive for SP/calretinin but immunonegative for CGRP. SP released from submucosal type II neurons probably acts via NK1Rs on type II and non-type II submucosal neurons to mediate intrinsic reflexes. EP3R-positive submucosal type II neurons may be potential targets of prostaglandin E2.
  • Distinct effects of CGRP on typical and atypical smooth muscle cells involved in generating spontaneous contractions in the mouse renal pelvis, Hikaru Hashitani, Richard J. Lang, Retsu Mitsui, Yoshio Mabuchi, Hikaru Suzuki, BRITISH JOURNAL OF PHARMACOLOGY, 158, (8) 2030 - 2045,   2009年12月, Background and purpose: We investigated the cellular mechanisms underlying spontaneous contractions in the mouse renal pelvis, regulated by calcitonin gene-related peptide (CGRP). Experimental approach: Spontaneous contractions, action potentials and Ca2+ transients in typical and atypical smooth muscle cells (TSMCs and ATSMCs) within the renal pelvis wall were recorded separately using tension and intracellular microelectrode recording techniques and Fluo-4 Ca2+ imaging. Immunohistochemical and electron microscopic studies were also carried out. Key results: Bundles of CGRP containing transient receptor potential cation channel, subfamily V, member 1-positive sensory nerves were situated near both TSMCs and ATSMCs. Nerve stimulation reduced the frequency but augmented the amplitude and duration of spontaneous phasic contractions, action potentials and Ca2+ transients in TSMCs. CGRP and agents increasing internal cyclic adenosine monophosphate (cAMP) mimicked the nerve-mediated modulation of TSMC activity and suppressed ATSMCs Ca2+ transients. Membrane hyperpolarization induced by CGRP or cAMP stimulators was blocked by glibenclamide, while their negative chronotropic effects were less affected. Glibenclamide enhanced TSMC Ca2+ transients but inhibited ATSMC Ca2+ transients, while both 5-hydroxydecanoate and diazoxide, a blocker and opener of mitochondrial ATP-sensitive K+ channels, respectively, reduced the Ca2+ transient frequency in both TSMCs and ATSMCs. Inhibition of mitochondrial function blocked ATSMCs Ca2+ transients and inhibited spontaneous excitation of TSMCs. Conclusions and implications: The negative chronotropic effects of CGRP result primarily from suppression of ATSMC Ca2+ transients rather than opening of plasmalemmal ATP-sensitive K+ channels in TSMCs. The positive inotropic effects of CGRP may derive from activation of TSMC L-type Ca2+ channels. Mitochondrial Ca2+ handling in ATSMCs also plays a critical role in generating Ca2+ transients.
  • Characterisation of calcitonin gene-related peptide-immunoreactive neurons in the myenteric plexus of rat colon, Retsu Mitsui, CELL AND TISSUE RESEARCH, 337, (1) 37 - 43,   2009年07月, A mechanical or chemical stimulus applied to the intestinal mucosa induces motility reflexes in the rat colon. Enteric neurons containing calcitonin gene-related peptide (CGRP) have been suggested as intrinsic primary afferent neurons responsible for mediating such reflexes. In the present study, immunohistochemistry was performed on whole-mount stretch preparations to investigate chemical profiles, morphological characteristics and projections of CGRP-containing neurons in the myenteric plexus of the rat colon. CGRP-positive neuronal cell bodies were detected in preparations incubated with colchicine-containing medium, whereas CGRP-positive nerve fibres were found in colchicine-untreated preparations. These neurons had large oval or round cell bodies that were also immunoreactive for the calcium-binding protein calretinin and neurofilament 200. Myenteric neurons positive for both calretinin and neurofilament 200 had several long processes that emerged from the cell body, consistent with Dogiel type II morphology. Application of the neural tracer DiI to the intestinal mucosa revealed that DiI-labelled myenteric neurons each had an oval or round cell body immunoreactive for calretinin. Thus, CGRP-containing myenteric neurons are Dogiel type II neurons and are immunoreactive for calretinin and neurofilament 200 in the rat colon. These neurons probably project to the intestinal mucosa.
  • Fibre-free diet leads to impairment of neuronally mediated muscle contractile response in rat distal colon, R. Mitsui, S. -I. Karaki, Y. Kubo, Y. Sugiura, A. Kuwahara, NEUROGASTROENTEROLOGY AND MOTILITY, 18, (12) 1093 - 1101,   2006年12月, Dietary fibre consumption is known to be beneficial to increase stool bulk and frequency. In contrast, it is unclear whether chronic dietary fibre deficiency affects colonic motor functions, especially neuronally mediated muscle contractions. In this study, rats were fed a fibre-free diet or diet containing dietary fibre (cellulose or guar gum) for 27 days. Furthermore, neurogenic and myogenic contractions were evaluated in circular and longitudinal muscle strips of the distal colon. Additionally, the number of enterochromaffin (EC) cells, which play important roles in the initiation of the peristaltic reflex, was also examined by immunohistochemistry for serotonin. Myogenic contractions induced by carbachol or substance P were examined in the presence of tetrodotoxin. Circular muscle was hyposensitive to carbachol, but longitudinal muscle was hypersensitive to substance P in the fibre-free group. Nerve-mediated circular (5-20 Hz) and longitudinal (1-2 Hz) muscle contractions evoked by electrical field stimulation were attenuated in the fibre-free group and the latter response was almost abolished by atropine, suggesting functional changes of cholinergic neurons. EC cell number was decreased in the fibre-free group. In conclusion, changes in neurogenic and myogenic contractions and a decrease in EC cell number observed may affect colonic motility of the fibre-free group.
  • Short-chain fatty acid receptor, GPR43, is expressed by enteroendocrine cells and mucosal mast cells in rat intestine, S Karaki, R Mitsui, H Hayashi, Kato, I, H Sugiya, T Iwanaga, JB Furness, A Kuwahara, CELL AND TISSUE RESEARCH, 324, (3) 353 - 360,   2006年06月, Short-chain fatty acids (SCFAs), such as acetate, propionate, and butyrate, are the major anions in the large intestinal lumen. They are produced from dietary fiber by bacterial fermentation and are known to have a variety of physiological and pathophysiological effects on the intestine. In the present study, we investigated the expression of the SCFA receptor, GPR43, in the rat distal ileum and colon. Expression of GPR43 was detected by reverse transcriptase/polymerase chain reaction (RT-PCR), Western blotting, and immunohistochemistry. mRNA for GPR43 was detected, by RT-PCR, in extracts of the whole wall and separated mucosa from the ileum and colon and from muscle plus submucosa from the ileum, but not from muscle plus submucosa preparations from the colon. We raised a rabbit antiserum against a synthesized fragment of rat GPR43; this was specific for rat GPR43. GPR43 protein was detected by Western blot analysis in extracts of whole wall and separated mucosa, but not in muscle plus submucosa extracts. By immunohistochemistry, GPR43 immunoreactivity was localized to enteroendocrine cells expressing peptide YY (PYY), whereas 5-hydroxytryptamine (5-HT)-immunoreactive (IR) enteroendocrine cells were not immunoreactive for GPR43. Mast cells of the lamina propria expressing 5-HT were also GPR43-IR. The results of the present study suggest that the PYY-containing enteroendocrine cells and 5-HT-containing mucosal mast cells sense SCFAs via the GPR43 receptor. This is consistent with physiological data showing that SCFAs stimulate the release of PYY and 5-HT from the ileum and colon.
  • Propionate modulates spontaneous contractions via enteric nerves and prostaglandin release in the rat distal colon, R Mitsui, S Ono, SI Karaki, A Kuwahara, JAPANESE JOURNAL OF PHYSIOLOGY, 55, (6) 331 - 338,   2005年12月, Short-chain fatty acids, such as propionate and acetate, are produced by a bacterial fermentation of carbohydrates in the colonic lumen. We examined the effects of propionate on the frequency and mean amplitude of spontaneous giant contractions (GCs) in circular muscle strips of the rat distal colon with the mucosa attached. An addition of propionate increased the frequency of GCs for about 20 min (>= 1 mm), but the mean amplitude was decreased ( >= 0.1 mm). The propionate-induced increase in the frequency of GCs was blocked by the muscarinic acetylcholine receptor antagonist, atropine. In contrast, the nicotinic receptor antagonist, hexamethonium, augmented the response. The propionate-induced decrease in the mean amplitude of GCs was prevented by the cyclooxygenase inhibitor, piroxicam. A pretreatment of the tissues with acetate prevented the propionate-induced modulations of the frequency and amplitude of GCs. These results suggest that propionate increases the frequency of GCs by an activation of cholinergic motor neurons and decreases the mean amplitude by a prostaglandin release. Propionate as well as acetate may be involved in the regulation of spontaneous circular muscle activity in the rat distal colon.
  • Muscarinic and 5-HT4 receptors participate in the regulation of the frequency of spontaneous contractions of the longitudinal muscle in rat distal colon, S Ono, R Mitsui, S Karaki, A Kuwahara, BIOMEDICAL RESEARCH-TOKYO, 26, (4) 173 - 177,   2005年08月, Spontaneous contractions of the intestine are thought to play an important role in the gastrointestinal motility, including peristalsis. In the present study, we investigated mechanisms for regulation of the frequency of spontaneous contractions, using longitudinal muscle strips in rat distal colon. Atropine significantly decreased the frequency of spontaneous contractions, indicating that neuromuscular transmission via muscarinic receptors increases the frequency of spontaneous contractions. SB-204070, 5-HT4 receptor antagonist also significantly decreased the frequency of spontaneous contractions, indicating that the activation of 5-HT4 receptors also increases the frequency of spontaneous contractions. In conclusion, it is suggested that muscarinic and 5-HT4 receptors participate in the regulation of the frequency of spontaneous contractions in the longitudinal muscle in rat distal colon, and that the frequency of spontaneous contraction is controlled by the enteric neurons.
  • Neural and non-neural mediation of propionate-induced contractile responses in the rat distal colon, R Mitsui, S Ono, S Karaki, A Kuwahara, NEUROGASTROENTEROLOGY AND MOTILITY, 17, (4) 585 - 594,   2005年08月, Short-chain fatty acids (SCFAs), including propionate, butyrate and acetate, are fermentation products of carbohydrates in the colon. We investigated the contractile effects of SCFAs on the rat distal colon. Mechanical activity of the circular muscle in strip preparations was recorded in vitro. Propionate and butyrate concentration-dependently (10 mu mol L-1-10 mmol L-1) induced rapid, large amplitude phasic contractions (the first phase) followed by tonic contractions (the second phase). Acetate itself had no effect on muscle activity, although preincubation with acetate attenuated both phases of the propionate-induced response. The propionate-induced phasic contraction was attenuated by atropine, tetrodotoxin and the 5-HT4 receptor antagonist SB-204070. The propionate-induced tonic contraction was attenuated by the cyclo-oxygenase inhibitor piroxicam. Antagonists of 5-HT1A, 5-HT2A and 5-HT3 receptors had no effect on the responses. Propionate-induced responses were not observed in mucosa-free preparations. These results suggest that propionate acts on receptors in the mucosa causing the release of 5-HT from enterochromaffin cells. 5-HT acts through 5-HT4 receptors on the endings of intrinsic primary afferent neurones that in turn activate cholinergic motor neurones that contract the circular muscle. Propionate also causes tonic contraction, via prostaglandin release, in the rat distal colon.
  • Distribution and ultrastructure of interstitial cells of Cajal in the gastric antrum of wild-type and Ws/Ws rats, R Mitsui, T Komuro, ANATOMY AND EMBRYOLOGY, 206, (6) 453 - 460,   2003年05月, Interstitial cells of Cajal (ICC) in the stomach of wild-type and Ws/Ws mutant rats that are deficient in c-kit were studied by immunohistochemistry and electron microscopy to elucidate their regional specialization in the gastric antrum. Immunohistochemistry for Kit protein demonstrated that in wild-type rats ICC were located at the submucosal border of the circular muscle layer (ICC-SM) in a limited extension of the antrum, from the pyloric sphincter towards the corpus, as well as within both the circular (ICC-CM) and longitudinal (ICC-LM) muscle layers and in the myenteric plexus region (ICC-AP). In c-kit mutant Ws/Ws rats while ICC-CM and ICC-LM were not observed, but unexpectedly, a few ICC-SM and ICC-AP were found. By electron microscopy, ICC-SM and ICC-AP were characterized by abundant mitochondria, many caveolae, a distinct basal lamina and formed gap junctions with other ICC or with smooth muscle cells and make close contacts with nerves. Thus, ICC-SM and ICC-AP of the rat antrum were classified as Type 3 ICC, the type most similar to smooth muscle cells. The functional significance of ICC-SM and their survival in the c-kit mutant animals is discussed in reference to the role of the c-kit/stem cell factor system for their cellular maturation.
  • Direct and indirect innervation of smooth muscle cells of rat stomach, with special reference to the interstitial cells of Cajal, R Mitsui, T Komuro, CELL AND TISSUE RESEARCH, 309, (2) 219 - 227,   2002年08月, Interstitial cells of Cajal in the circular (ICCCM) and longitudinal (ICC-LM) muscle layer of the rat gastric antrum and their innervation were studied ultrastructurally. Both ICC-CM and ICC-LM are characterized by many mitochondria, rough and smooth endoplasmic reticulum, caveolae, and formation of gap junctions with each other and with muscle cells, though ICC-LM tend to show more variable cytoplasmic features depending on section profiles. Close contacts between nerve terminals and both ICC-CM and ICC-LM are observed. These possible synaptic structures are characterized by: (1) accumulation of synaptic vesicles in nerve varicosities, (2) a narrow gap (about 20 nm) between pre- and postjunctional membranes, (3) lack of a basal lamina between pre- and postjunctional membranes, and (4) the presence of an electron-dense lining on the inner aspect of prejunctional membranes. Almost the same characteristics are observed between the nerve terminals and the muscle cells of both circular and longitudinal muscle layers of the same specimens. Therefore, we conclude that the smooth muscle cells of both circular and longitudinal layers of the rat antrum are directly and indirectly innervated via ICC. Their functional significance is discussed.

書籍等出版物

  • Role of pericytes in the initiation and propagation of spontaneous activity in the microvasculature. In: Advances in Experimental Medicine and Biology, Smooth muscle spontaneous activity., Hashitani H, Mitsui R, Springer,   2019年

受賞

  •   2018年08月, 日本平滑筋学会, 日本平滑筋学会第4回白鳥常男賞


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