* * Fischer 344 carbachol carbachol M phenylephrine - phenylephrine [ 3 H] quinuclidinyl benzilate (QNB) [ 3 H] QNB QOL quality of life 1 3) 2
Krebs-Henseleit 37 0.1 10 ml organ bath 3 7T-15-240 3 mm NaCl 118, KCl 4.7 NaHCO 3 24.9 MgSO 4 1.2 KH 2 PO 4 1.2 glucose 11.1 ascorbic acid 0.057 CaCl 2 1.8 ph 7.4 organ bath 3 KCl 66.7mM KCl K + camp K + 37 3), 4 propranolol 10-6 M carbachol 10-10 M 3 10-5 M carbachol atropine 10-7 Mcarbachol Fischer 344 5 g Tris-HCl buffer 25mM Tris, 10mM MgCl 2,. Krebs-Henseleit NaCl 56 KCl 66.7mM prazosin 10-7 M pirenzepine M 10-7 M 10-6 M methoctramine M 10-7 M 10-6 M p-f-hhsid M 10-7 M 10-6 M carbachol Schild plot phenylephrine 10-8 M 3 10-4 M. ph 7.4 7.5 ml mm Kinematica, Luzern, Switzerland 95%O2, 5% CO2
Lowry 6 1mg/ml [ 3 H] quinuclidinyl benzilate QNB carbamylcholine chloride phenylephrine Whatman GF/C M24R, Brandel, Gaitherberg, MD, USA tetrahydrochloride Sigma Chemical Co. St. Louis, MO, USA atropine sulfate mono- hydrate mltris-hcl buffer [ H] QNB [ 3 H]QNB New England Nuclear Bos- ton, MA, USA II, Amersham, Arlington Heights, IL, USA ml ACS- Tri- Carb 2300TR, Liquid Scintillation Analyzer, Packard, Downers Grove, IL, USA atropine 10 6 M Scatchard plot QNB hydrochloride prazosin hydrochloride pro- Tris-HCl buffer ml pranolol hydrochloride hexahydro-sila-difenidolhydrochloride p-fluoroanalog (p-f-hhsid) pirenzepine hydrochloride methoctramine Student s t-test Welch s t-test + N.S. not significant carbachol K + % carbachol% carbachol carbachol atropine10-7 M carbachol atropine10-7 M
carbachol pirenzepine A K + % carbachol% carbachol carbachol pirenzepine 10-7 Mpirenzepine 3 10-7 M pirenzepine 10-6 M B carbachol pirenzepine Schild plot dose ratio carbachol p-f-hhsid A K + % carbachol% carbachol carbachol p-f-hhsid 10-7 Mp-F-HHSiD 3 10-7 M p-f-hhsid 10-6 M B carbachol p-f-hhsid Schild plot dose ratio
P<0.05. K + 2.8 0.1 g, n 8, 3.3 0.2 g, n 6 2.2 0.3 g, n 8, 2.6 0.3 g, n 6 K + carbachol phenylephrine K + carbachol carbachol methoctramine K + % 127.8 14.4% n 8 carbachol% 157.0 13.8% n 6 EC 50 carbachol (6.82 2.57) 10-7 M (9.08 1.57) carbacholmethoctramine10-7 M methoctramine 3 10-7 M 10-7 M methoctramine 10-6 M carbachol 3 H QNB 3 H QNB A B
atropine carbachol 4 nm pirenzepine, p-f-hhsid carbachol Schild plot pirenzepine 0.26 0.05, n=4, 0.22 0.04, n=4) 0.87 p-f-hhsid0.98 carbachol ) p-f-hhsid pa2 7.42 0.38 methoctramine phenylephrine 2), 7 58.9 6.4%, n 8, 86.3 4.3%, n 6, P<0.01 EC 50 (4.65 0.94) 10-6 M, (4.31 0.73) 10-6 M phenylephrine K + % phenylephrine% phenylephrine *P 0.05 **P 0.01(vs ). [ 3 H]QNB 0.25 Bmax fmol / mg protein 50.7 3.4, n=4, 49.8 4.5, n=4 Kd ( (, Ca 2+ Ca 2+ KCl KCl 66.7mM K + Ca 2+ Young Aged Bmax (fmol/mg protein) 50.7 3.4 49.8 4.5 Kd(nM) 0.26 0.05 0.22 0.04 Bmax Kdn n 4 4
Ordway 8 KCl 3 Nishimoto 4 Saito 9 KCl Ca 2+ Fischer344 K + carbachol phenylephrine phenylephrine, norepinephrine, clonidine EC 50 carbachol EC 50 3 13 Ordway 8 7 Fischer 344 acetylcholine, bethanechol, oxotremorine EC 50 Sprague-Dawley 10 Fischer 344 11 Fischer 344 Hayes 15 Sprague-Dawley Nishimoto 3), 4 Ordway 12 Bmax fmol/mg protein Kd Kolta 14 phenylephrine
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Yamagata Med J 2004 22 (2) 129-138 Tomomi Aita, Kozue Aita, Akiko Yamada, Akira Ishihata, Yumi Katano Division of Clinical Nursing, Department of cardiovascular Physiology, Yamagata University School of Medicine, Yamagata, Japan ABSTRACT The urinary bladder is innervated and functionally regulated by the autonomic nervous system. In order to elucidate the mechanisms of age-related changes in urinary bladder function, we studied the influence of aging on, 1) the contractile response to carbachol in the body and to phenylephrine in the trigone, 2) the binding characteristics of [ 3 H] quinuclidinyl benzilate (QNB) to muscarinic cholinoceptors in rat urinary bladder. Urinary bladder from young (2-3 months) or aged (27 months) male Fischer 344 rat was isolated. Carbachol-induced contraction was not significantly changed by aging. The contraction was competitively antagonized by p-f-hhsid (M 3-blocker). In contrast, phenylephrine elicited greater contraction in aged trigone than that in young rat. In the receptor assay, [ 3 H] QNB was used as a muscarinic receptor ligand. [ 3 H] QNB binding characteristics were not significantly changed by aging. Because the -adrenoceptor-mediated relaxation was reportedly attenuated in aged bladder, the facilitation of -receptor-mediated contraction and the decreased -receptor-mediated relaxation may cause voiding difficulty in the aged urinary bladder. Key words : bladder, smooth muscle, aging, muscarinic cholinoceptors, -adrenoceptors