Investigation of neurotransmission in vas deferens from alpha(2A/D)-adrenoceptor knockout mice

Abstract

1. We have investigated pre- and post-junctional responsiveness in vas deferens from wild-type and alpha(2A/D)-adrenoceptor knockout mice. The response to a single stimulus was not significantly different between wild-type and knock-out mice. The isometric contraction to 10 Hz stimulation for 4 s was significantly larger in vas deferens from knockout as compared with wild-type. 2. The maximum potentiation of 10 Hz stimulation-evoked contractions by yohimbine was to 206.2+/-38.0% of control in wild-type but to 135.8+/-13.6% of control in knockout. The alpha(2A/D)-adrenoceptor selective antagonist BRL 44408 significantly increased the 10 Hz stimulation-evoked contraction in wild-type but not knockout, and the reverse was true for the alpha(2C)-adrenoceptor selective antagonist spiroxatrine. The alpha(2B)-adrenoceptor antagonist imiloxan had no effect on the evoked contraction except at high concentrations, and only in wild-type. Following cocaine (3 microM) and BRL 44408 (1 microM), 10 Hz responses were similar in shape and maximum between wild-type and knock-out. 3. The alpha(2)-adrenoceptor agonist xylazine virtually abolished the early component of the contraction to 10 Hz stimulation in the presence of nifedipine (10 microM) in vas deferens from knockout mice in a way consistent with a change of receptor subtype but without clear evidence for a reduced receptor number. However, the late component of the contraction to 10 Hz stimulation was significantly potentiated by xylazine in tissues from knock-out mice. 4. It is concluded that, although non-alpha(2A/D)-adrenoceptors replace alpha(2D)-adrenoceptors in this knockout, the alpha(2)-adrenoceptor agonist and antagonist data are contradictory. The antagonist data suggest a major loss of prejunctional alpha(2)-adrenoceptors, but this is not necessarily supported by the agonist data.

Figure 1

Typical isometric contractions to 10 Hz stimulation for 4 s in vas deferens from wild-type (WT) and α_2A/D-adrenoceptor knockout (ko) mice. (a) Control response and response following cocaine (3 μM) and subsequent BRL 44408 (1 μM). (b) Response in the presence of nifedipine (10 μM). Time and tension scales have arbitrary zero points.

Figure 2

Effects of the α₂-adrenoceptor antagonists yohimbine (yoh, non-selective), BRL 44408 (BRL, α_2D-selective), imiloxan (imil, α_2B-selective) and spiroxatrine (spir, α_2C-selective) on isometric contractions evoked by 10 Hz electrical stimulation for 4 s in vas deferens from wild-type (a) or α_2A/D-adrenoceptor knockout (b) mice. Vertical bars indicate s.e. of mean from four (BRL 44408, imiloxan) or six (other antagonists) experiments.

Figure 3

Effects of cocaine (3 μM) and the α₂-adrenoceptor antagonist BRL 44408 (1 μM) on isometric contractions evoked by 10 Hz electrical stimulation for 4 s in vas deferens from wild-type or α_2A/D-adrenoceptor knockout mice. Vertical bars indicate s.e. of mean from at least five experiments. Asterisks denote significance of difference from control responses (^* P<0.05).

Figure 4

Effects of the α₂-adrenoceptor agonist xylazine, in the presence of vehicle (veh) or the α₂-adrenoceptor antagonist BRL 44408 1 μM (BRL, α_2D-selective), on isometric contractions evoked by 10 Hz electrical stimulation for 4 s in vas deferens from wild-type (WT) or α_2A/D-adrenoceptor knockout (ko) mice in the presence of cocaine (3 μM). Vertical bars indicate s.e. of mean from at least four experiments.

Figure 5

Effects of the α₂-adrenoceptor agonist xylazine, in the presence of vehicle on isometric contractions evoked by 10 Hz electrical stimulation for 4 s in vas deferens from wild-type (WT) or α_2A/D-adrenoceptor knockout (ko) mice in the presence of cocaine (3 μM). Effects of xylazine 0.01–1 μM (WT) or 0.1–10 μM (ko) on contractions are shown at various times (0.5, 1, 2, 3, 4 s) from beginning of 10 Hz stimulation. Note that xylazine inhibits at all time points in WT, but inhibits at 0.5–1 s and potentiates at 2–4 s in ko (at least with 1 and 10 μM). Vertical bars indicate s.e. of mean from at least four experiments. The insets show typical isometric responses to 10 Hz stimulation before and after xylazine (1 or 10 μM) in tissues from a knockout (ko) and a wild-type (WT) mouse. Time and tension scales have arbitrary zero points.

Figure 6

Effects of the α₂-adrenoceptor agonist xylazine, in the presence of vehicle (veh), the α₂-adrenoceptor antagonist BRL 44408 1 μM (BRL, α_2D-selective) or the α₁-adrenoceptor antagonist prazosin (praz), on isometric contractions evoked by 10 Hz electrical stimulation for 4 s in vas deferens from wild-type (WT) or α_2A/D-adrenoceptor knockout (ko) mice in the presence of nifedipine (10 μM). Vertical bars indicate s.e. of mean from at least four experiments. Effects of xylazine in the presence of vehicle are taken from Rajamani et al. (2001).