Investigation of postjunctional alpha1- and alpha2-adrenoceptor subtypes in vas deferens from wild-type and alpha(2A/D)-adrenoceptor knockout mice

Abstract

1. The subtypes of alpha(1)- and alpha(2)-adrenoceptor mediating contractions of vas deferens have been examined in wild-type and alpha(2A/D)-adrenoceptor knockout mice. 2. Maximum contractions to noradrenaline but not phenylephrine were significantly greater in vas from wild-type. The alpha(1A)-adrenoceptor antagonist RS100329 (5-methyl-3-[3-[4-[2-(2,2,2,-trifluoroethoxy)phenyl]-1-piperazinyl]propyl]-2,4-(1H)-pyrimidinedione) (10 nM) significantly shifted the potency of noradrenaline. The alpha(2D)-adrenoceptor antagonist BRL 44408 (2-[(4,5-dihydro-1H-imidazol-2-yl)methyl]-2,3-dihydro-1-methyl-1H-isoindole) significantly reduced the maximum contraction to noradrenaline in wild-type but not in knockout. 3. Following prazosin (0.1 micro M), a component of the contraction to noradrenaline in wild-type but not in knockout was sensitive to the alpha(2)-adrenoceptor antagonist yohimbine. 4. Nifedipine (10 micro M) or suramin (100 micro M) reduced the contraction to 10 Hz stimulation for 4 s to an early peak and small maintained response. The peak was abolished by the alpha(1)-adrenoceptor antagonist prazosin. 5. RS100329 or prazosin inhibited 10 Hz stimulation evoked contractions with a U-shaped concentration-response curve: inhibiting responses up to 0.1 micro M, with a reversal of inhibition above this concentration. In the presence of suramin or nifedipine, the reversal of inhibition by high concentrations of prazosin was reduced. 6. The alpha(1D)-adrenoceptor selective antagonist BMY7378 (8-[2-(4-(2- methoxyphenyl)piperazin-1-yl)ethyl]-8-azaspiro[4,5]decane-7,9-dione) produced inhibition of 10 Hz evoked contractions only in high concentrations. 7. In conclusion, contractions to nerve stimulation in mouse vas deferens involve largely alpha(1A)-adrenoceptors and purinoceptors. alpha(1)-Adrenoceptor antagonists in high concentrations increase the purinergic response presumably by blocking prejunctional alpha(2)-adrenoceptor-mediated inhibition. In the presence of nifedipine, responses are predominantly alpha(1)-adrenoceptor mediated. Contractions to exogenous noradrenaline involved both alpha(1A)- and alpha(2A/D)-adrenoceptors in wild-type mice.

Figure 1

Concentration–response curves to noradrenaline following vehicle (veh) or BRL 44408 (1 μM) (BRL) in vas deferens from wild-type (WT) and α_2A/D-adrenoceptor knockout mice (ko) in the first series of experiments. The dashed lines indicate the maximum tensions obtained (sum of maximum tensions obtained in each individual tissue irrespective of concentration at which they were obtained). Vertical bars indicate s.e.m. from seven to 22 animals. Maximum responses to noradrenaline following vehicle in wild-type were significantly greater than in knockout. BRL 44408 significantly decreased the maximum contraction to noradrenaline only in wild-type. The inset shows a typical tension recording in a tissue from a wild-type animal, in which a maximum response is obtained to 10 μM, with subsequent decline (responses were qualitatively similar in tissues from knockout); tension and time scales have an arbitrary zero point.

Figure 2

Concentration–response curves to noradrenaline following prazosin (0.1 μM) (praz) or prazosin (0.1 μM)/yohimbine (1 μM) (praz/yoh) in vas deferens from wild-type (WT) and α_2A/D-adrenoceptor knockout mice (ko) in the second series of experiments. Vertical bars indicate s.e.m. from eight to 11 animals. The response to noradrenaline (10 μM) following prazosin in wild-type was significantly larger than in the other three groups.

Figure 3

Effects of prazosin (praz) or RS 100329 (RS) on isometric contractions evoked by field stimulation at 10 Hz for 4 s in vas deferens from wild-type (WT) and knockout mice (ko): (a) in the presence of vehicle (veh), or (b) in the presence of suramin (100 μM) (sur) or nifedipine (10 μM) (nif). Vertical bars indicate s.e.m. from three to 11 animals.

Figure 4

Typical recordings of contractions evoked by 10-Hz stimulation for 4 s in vas from knockout mice: (a) control, (b) following suramin (100 μM) or (c) following nifedipine (10 μM) (qualitatively similar in tissues from wild-type). Tension and time scales have arbitrary zero points.

Figure 5

Effects of BMY 7378 on isometric contractions evoked by field stimulation at 10 Hz for 4 s in the absence (veh) or presence of nifedipine (10 μM) (nif) in vas deferens from wild-type (WT) or knockout mice (ko). Vertical bars indicate s.e.m. from six animals.