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Calcitonin gene-related peptide (CGRP) binding sites in the female guinea pig genital tract.
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CALCITONIN GENE-RELATED PEPTIDE (CGRP) BINDING SITES IN THE FEMALE GUINEA PIG GENITAL TRACT.
R.F. POWER,A.J. RUTHERFORD 1, S.P. SALAS2, J. WHARTON,J.M. POLAK. Departments of Histochemistry, 1Obstetrics and 2Medicine, Royal Postgraduate Medical School, Hammersmith Hospital, Du Cane Road, London W12 ONN. In the present study, we have investigated the distribution of CGRPbinding sites in the genital tract of the female guinea pig using the technique of in v i t r o receptor autoradiography. Adult female Dunkin Hartley guinea pigs - T6~Twere k i l l e d by an overdose of ether, the genital tract rapidly removed, dissected into d i f f e r e n t regions and snap frozen. Cryostat sections (lO#m) were incubated with 1251-alpha CGRP (lO-200pm) for 30 minutes at 25"C and autoradiograms generated by exposing l~belled sections to radiation-sensitive photographic film (A~rsham Hyperfilm-~H) for 6-12 days at 4°C, To determine non-specific binding serial sections were incubated with the radioligand in the presence of an excess of unlabelled CGRP (lO-IM). Specific binding sites were i d e n t i f i e d on the epithelium and smooth muscle of the vagina, cervix, corpus uteri and fallopian tube but there was an apparent absence of binding sites in the ovary. In addition, in keeping with the known vasodilator effect of CGRP on the peripheral circulation, blood vessels (arteries) in all tissues examined were also heavily labelled. CGRP-immunoreactivenerve fibres have been described in the genital tract of some species, including guinea pig, and CGRP is known to i n h i b i t spontaneous uterine contractions. This localization of specific binding sites for CGRP suggests a direct effect on smooth muscle and also indicates a possible role for CGRP in local control of uterine blood flow. RFP is a WeIIcome Trust Research Fellow. SPS is a Fellow of ODEPLAN, Chile.
SHEAR STRESS CAUSESRELEASE OF SUBSTANCE P FROMRAT HINDLIMB VASCULAR ENDOTHELIAL CELLS V. RALEVlC, P. MILNER, O. HUDLICKA, F. KRISTEK and G. BURNSTOCK. Department of Anatomy and Developmental Biology, University College London, Gower Street, London WCIE 6BT. A three-fold increase in perfusion rate of Krebs' solution containing BSA through the rat hindlimb vasculature to create shear stress caused a s i g n i f i c a n t release of substance P (4.4 ± 3.4 to 30.7 ± 9.5 pmol/min, P(O.05) which was reproduced 20 min later in the same animal (I.6 ± 0.5 to 37.8 ± 6.7 pmol/min, P(O.OOl, n = 5). Destruction of primary sensory neurones by capsaicin did not reduce the extent of substance P release on either the f i r s t or second period of high flow (2.7 ± 1.5 to 44.9 ± 13.6 pmol/min, P(O.02 followed by 4.4 ± 1.8 to 41.8 ± l l . 9 pmol/min, P(O.02, n = 5). However, after removal of the endothelium by a i r treatment (a check with electronmicroscopy showed loss of endothelial cells in arteries, but not arterioles or c a p i l l a r i e s ) between the two periods of high flow, there was no significant release of substance P (2.2 ± 0.6 to 7.2 ± 4.1 pmol/min, n = 5). We conclude that high flow causing shear stress evokes release of substance P largely from endothelial cells in the arterial tree.
CALCITONIN GENE-RELATED PEPTIDE (CGRP) BINDING SITES IN THE FEMALE GUINEA PIG GENITAL TRACT.
R.F. POWER,A.J. RUTHERFORD 1, S.P. SALAS2, J. WHARTON,J.M. POLAK. Departments of Histochemistry, 1Obstetrics and 2Medicine, Royal Postgraduate Medical School, Hammersmith Hospital, Du Cane Road, London W12 ONN. In the present study, we have investigated the distribution of CGRPbinding sites in the genital tract of the female guinea pig using the technique of in v i t r o receptor autoradiography. Adult female Dunkin Hartley guinea pigs - T6~Twere k i l l e d by an overdose of ether, the genital tract rapidly removed, dissected into d i f f e r e n t regions and snap frozen. Cryostat sections (lO#m) were incubated with 1251-alpha CGRP (lO-200pm) for 30 minutes at 25"C and autoradiograms generated by exposing l~belled sections to radiation-sensitive photographic film (A~rsham Hyperfilm-~H) for 6-12 days at 4°C, To determine non-specific binding serial sections were incubated with the radioligand in the presence of an excess of unlabelled CGRP (lO-IM). Specific binding sites were i d e n t i f i e d on the epithelium and smooth muscle of the vagina, cervix, corpus uteri and fallopian tube but there was an apparent absence of binding sites in the ovary. In addition, in keeping with the known vasodilator effect of CGRP on the peripheral circulation, blood vessels (arteries) in all tissues examined were also heavily labelled. CGRP-immunoreactivenerve fibres have been described in the genital tract of some species, including guinea pig, and CGRP is known to i n h i b i t spontaneous uterine contractions. This localization of specific binding sites for CGRP suggests a direct effect on smooth muscle and also indicates a possible role for CGRP in local control of uterine blood flow. RFP is a WeIIcome Trust Research Fellow. SPS is a Fellow of ODEPLAN, Chile.
SHEAR STRESS CAUSESRELEASE OF SUBSTANCE P FROMRAT HINDLIMB VASCULAR ENDOTHELIAL CELLS V. RALEVlC, P. MILNER, O. HUDLICKA, F. KRISTEK and G. BURNSTOCK. Department of Anatomy and Developmental Biology, University College London, Gower Street, London WCIE 6BT. A three-fold increase in perfusion rate of Krebs' solution containing BSA through the rat hindlimb vasculature to create shear stress caused a s i g n i f i c a n t release of substance P (4.4 ± 3.4 to 30.7 ± 9.5 pmol/min, P(O.05) which was reproduced 20 min later in the same animal (I.6 ± 0.5 to 37.8 ± 6.7 pmol/min, P(O.OOl, n = 5). Destruction of primary sensory neurones by capsaicin did not reduce the extent of substance P release on either the f i r s t or second period of high flow (2.7 ± 1.5 to 44.9 ± 13.6 pmol/min, P(O.02 followed by 4.4 ± 1.8 to 41.8 ± l l . 9 pmol/min, P(O.02, n = 5). However, after removal of the endothelium by a i r treatment (a check with electronmicroscopy showed loss of endothelial cells in arteries, but not arterioles or c a p i l l a r i e s ) between the two periods of high flow, there was no significant release of substance P (2.2 ± 0.6 to 7.2 ± 4.1 pmol/min, n = 5). We conclude that high flow causing shear stress evokes release of substance P largely from endothelial cells in the arterial tree.