While noradrenaline can cause contraction by increasing the release of Ca²⁺ from intracellular stores, it can also sensitise myofilaments to Ca²⁺. One pathway by which this sensitization can occur is the Rho-kinase pathway (Somlyo et al., 2000). Ca²⁺-sensitization can now be investigated in intact tissues with the recently-developed membrane permeable Rho kinase inhibitor (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamine dihyrochloride monohydrate (Y27632). It has been reported that Y27632 inhibits both the noradrenergic and purinergic components of contraction in the mouse vas deferens (Büyükafsar et al., 2003), based on the assumption that the first component of contraction is entirely purinergic. While this assumption is valid in other species, it is not the case for the mouse (Cleary et al., 2003). In the present study, the effects of Y27632 (3, 10, 30 and 100 µM) on pharmacologically-isolated noradrenergic and purinergic contractions of the mouse isolated vas deferens were investigated.

Male Balc/C mice, of weight 20 – 30 g, were killed by a Schedule 1 method (concussion and cervical dislocation). All statistical tests are Student’s T-tests (two-tailed; paired for contraction studies, unpaired for electrophysiology). In a preliminary study, the equilibrium time for the highly selective P2X1 receptor antagonist 4, 4’, 4’’, 4’’’-(carbonylbis(imino-5, 1, 3-benzenetriyl-bis(carbonylimino)))tetrakisbenzene-1, 3-disulphonic acid-octasodiumsalt (NF449; 10 µM) in contraction baths was 30 ± 5 min (n = 3). After pre-treatment with NF449 (10 µM), electrical field stimulation (EFS) induced contractions that were reduced in amplitude by 30 µM Y27632 (by 14 ± 5 %; n = 3; P < 0.05). In the presence of the
₁-adrenoceptor antagonist prazosin (100 nM) Y27632 did not significantly affect resting tone at any concentration (n = 3; P < 0.05,) and only decreased the amplitude of contraction induced by EFS at a concentration of 100 µM (17 ± 2%; n = 3; P < 0.05). Intracellular electrophysiological recordings showed that Y27632 (30 µM) had no detectable effect on purinergic transmission, as excitatory junction potential (EJP) amplitudes and fall times were not affected (n = 6; P > 0.05;). Similarly, there was no effect on the frequency of spontaneous EJPs (n = 6; P > 0.05;), suggesting that Y27632 did not interfere with ATP release.

In conclusion, Y27632 marginally reduces the noradrenergic component of phasic contraction. The effect of Y27632 on purinergic transmission only occurs at high concentrations and is argued to be a non-specific affect. There was no evidence for an effect of Y27632 on neurotransmitter release.

Büyükafsar K et al. (2003) Br J Pharmacol22, 1-7
Cleary, L. et al. (2003) Br J Pharmacol138, 1069-1076
Somlyo, A.P. et al. (2000) J Physiol522, 177-185

We gratefully acknowledge the provision and guidance provided by Dr T.C. Cunnane