Investigation of the role of 1D-adrenoceptors in contraction of mouse femoral small arteries using 1D-adrenoceptor-KO mice J. Zacharia1, C. Hillier1, C.J. Daly2, J.C. McGrath2, G Tsujimoto3, A. Tanoue3 & A. MacDonald1. Department of Biological and Biomedical Sciences, Glasgow Caledonian University, Glasgow, G4 0BA. 2Division of Neuroscience & Biological systems, IBLS, University of Glasgow, Glasgow, G12 8QQ. 3Department of Genomic Drug Discovery Science, Kyoto University, Japan.		Print Abstract Search PubMed for: Zacharia J Hillier C Daly CJ McGrath JC Tsujimoto G Tanoue A MacDonald A

The role of _1D-adrenoceptors in responses of mouse femoral small arteries to exogenous noradrenaline and electrical field stimulation (EFS) was examined using _1D-adrenoceptor-knockout (KO) mice. Antagonists prazosin (₁-selective), 5-methyl-urapidil (5MU, _1A-selective), BMY 7378 (_1D-selective) and the preferential _1B-alkylating agent, chloroethylclonidine (CEC) were used.

Second order branches of femoral artery (c. 175 µm) were dissected out from male 129/Sv/C57BL/6J (wild type, WT) and _1D-KO mice (25-32 g, 16 weeks old) and mounted on a small vessel wire myograph in physiological salt solution at 37^oC continuously bubbled with carbogen for isometric recording. For electrical field stimulation (EFS), vessels were incubated with 0.1µM RS 79948 (₂-adrenoceptor antagonist) and were stimulated at 20 V and 0.05 ms pulse width applied for 10 sec at frequencies of 2-20 Hz. Antagonist potency was expressed as the -log of the concentration required to inhibit the responses to EFS by 50% (pIC₅₀).

_1D-KO mice showed a 3-fold decrease in sensitivity to exogenous NA (pEC₅₀s: WT, 6.9±0.1;_1D-KO, 6.4±0.1, P<0.001, n=10). Prazosin and 5MU inhibited responses to exogenous NA and to nerve stimulation with high affinity with no significant differences between WT and _1D-KO. BMY 7378 (10-100 nM) had no effect on responses to NA in WT or _1D-KO. However, BMY 7378 (1 µM) produced significant shifts of the NA CRC with no significant difference in the pK_Bs (pK_Bs: WT, 7.1± 0.2; _1D-KO, 6.8±0.2, n=4). In WT nerve-mediated responses were more sensitive to BMY 7378 at lower frequencies (2 and 5 Hz) than at higher frequencies (e.g. pIC₅₀s: 5 Hz, 8.08±0.08; 10 Hz, 6.43±0.11, n=5, P<0.05) whereas in _1D-KO the sensitivity to BMY 7378 was the same at all frequencies (e.g. pIC₅₀s: 5 Hz, 6.43±0.06; 10 Hz 6.13±0.15, n=5, P>0.05). CEC (1 and 10 µM) produced a greater inhibition of exogenous NA in _1D-KO compared to WT (e.g. % of control maximum responses after 10 µM; WT, 79±2%, n=4; _1D-KO, 43±2%, n=3, P<0.01). CEC inhibited neurogenic responses in WT only at low frequencies (e.g. % of control, CEC, 10 µM, n=4; 5 Hz, 79±2, P<0.001; 10 Hz, 99±2, P>0.05) while in a1D-KO nerve-mediated responses were inhibited by CEC at all frequencies (e.g. % of control, CEC, 10 µM, n=7; 5 Hz, 71±6; 10 Hz, 84±4).

In conclusion, responses to exogenous noradrenaline and to nerve stimulation in mouse femoral small arteries are predominantly mediated by _1A-adrenoceptors. Loss in _1D-KO of the BMY 7378-sensitive component of the nerve-mediated responses at low frequencies of stimulation confirms that _1D-adrenoceptors contribute to responses to nerve stimulation. The increased sensitivity to CEC may suggest an up-regulation of _1B-adrenoceptors in _1D-KO mice.