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Detection of an alpha 2C adrenoceptor component in the alpha 2 agonist mediated inhibition of neurally evoked rat vasa deferentia contraction The α2A-adrenoceptor is thought to mediate α2 agonist induced inhibition of nerve-evoked contractions in rat vasa deferentia (Smith et al., 1992). Studies in knockout mice have also suggested the potential for α2B and/or α2C involvement, at least in the absence of α2A /D (Cleary L. et al, 2002; Trendelenburg et al., 2001, 2003). The aim of this study was to investigate whether α2C -adrenoceptor subtypes have any functional significance in ‘normal’ rat vasa deferentia contraction utilising a range of antagonists with varying selectivity for the α2C adrenoceptor. All experiments were carried out in compliance with UK legislation. Whole lengths of vasa deferentia were removed from euthanased, male, Sprague Dawley rats (250-400 g) and 2 tissue lengths strips (2x2x5 mm) dissected from the prostatic portion of each. Isometric tension recordings were performed with tissues bathed in Krebs’ solution (37°C, 95% O2 / 5% CO2) containing propranolol (1μM), doxazosin (0.1μM) and naproxen (10μM). Tissues were allowed to equilibrate (resting tension 0.5 g) for 1hr before electrical field stimulation (EFS, 0.2 ms, 20 Hz, 1 sec train, 25 volts) induced contractions were evoked at 1 minute intervals. The effect of UK14304 on EFS evoked contractions was investigated in the absence and presence of 5 concentrations of each antagonist. Where possible antagonist affinity (pKB) was calculated utilising Angus analysis (Lew & Angus, 1995) or an apparent pKB was calculated using the dose ratio at the lowest concentration. Data are reported as pKBs with 95% confidence intervals (CIs), N=5-8. CRCs for UK-14304 were shifted to the right by the α2-adrenoceptor specific antagonists yohimbine, idazoxan, efaroxan, RX821002 and RS79948 (pKB –8.0 (-7.9, -8.2), -8.2 (-8.0, -8.3), -8.3 (-8.2, -8.4), -8.9 (-8.7, -9.2) and –9.7 (-9.3, -10.0), respectively) and the α2A subtype preferring antagonists BRL44408 and BRL48962 pKB –7.4 (-7.1, -7.8) and -7.4 (-7.2, -7.5) respectively). The α2C selective antagonists, OPC28326, spiroxatrine, and prazosin produced an initial rightward shift at the lowest concentration, no further rightward shift was observed upon increasing concentrations, apparent pKBs were calculated using the dose ratio at the lowest concentration (pKB –7.7 (-6.7, -8.7), -8.3 (-7.7, -8.9) and –7.7 (-6.6, -8.8) respectively). The α2C preferring antagonists rauwolscine and MK912 produced rightward shifts but the Angus calculated pKBs were inconsistent with reported values at any α2-adrenoceptor subtype, therefore an apparent pKB was calculated at the lowest and highest concentrations utilised, the apparent pKB values agreed with reported values for the α2C and α2A-adrenoceptor subtype respectively. The α2B preferring antagonists ARC239 and the Juvantia compound (patent WO 03-008387; 10, 30, 50 100 and 300 nM) did not cause any significant shift in the UK-14304 CRC. This data suggests the role of the α2C-adrenoceptor subtype is frequency dependent and that independent blockade of the α2C and α2A -adrenoceptor subtype p roduces less contraction attenuation compared with blockade of both either by a non-selective antagonist; RS79948, or a α2C and α2A -adrenoceptor antagonist , OPC28326 and BRL44408 respectively , suggesting some synergy between the α2C and α2A -adrenoceptors. It is concluded that the α2C-adrenoceptor is functionally present on excitatory sympathetic nerve terminals of the vasa deferentia in normal rats and plays a small role in α2-adrenoceptor agonist induced inhibition of nerve-evoked contractions.
Smith et al. (1992). Eur. J. Pharmacol. 211, 251-256.
Patent WO 03/008387 30.01.2003 exemplified compound C. |
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