Statins restore KCa2.3 channel function in the rat middle cerebral artery

Alister McNeish1, Francesc Jimenz-Altayo2, Christopher Garland2. 1Department of Pharmacy and Pharmacology, University of Bath, Bath, BA27AY, United Kingdom, 2Department of Pharmacology, Oxford University, Oxford, OX13QT, United Kingdom.

Introduction In the rat middle cerebral artery inhibition of NOS or stimulation of TP leads to the loss of KCa2.3 function, subsequent inhibition of TP receptors or Rho kinase (ROCK) restores this component of hyperpolarization (McNeish et al 2007, McNeish et al BPS winter meeting 2010). We investigated if HMG-CoA inhibitors, statins (which are known to interfere with ROCK signalling via pleiotropic actions) could protect KCa2.3 function of endothelium dependent hyperpolarization (EDH) in vessels able to synthesize NO or restore KCa2.3 function to the isolated EDHF response of arteries treated with a NOS inhibitor.

Methods: Middle cerebral arteries from male Wistar rats (250-300g) were mounted on a wire myograph for tension recording. Sharp glass microelectrodes were impaled in smooth muscle cells for simultaneous recording of membrane potential as previously described (McNeish et al. 2007)

Results: In middle cerebral arteries able to synthesise NO but treated with the TP receptor agonist U46619 (100 nM) endothelium dependent hyperpolarization (EDH) evoked by the PAR2 agonist SLIGRL (20 µM; 30.9±5.9mV n = 7) was unaffected by simvastatin (100 nM; 31.8±4.4 mV, n = 7) or subsequent addition of the KCa3.1 inhibitor TRAM-34 (1 µM; 25.4±6.4 mV, n = 4); however additional inhibition of KCa2.3 with apamin significantly reduced EDH (15.8 ± 2.9 mV n = 4). In Vessels where NOS is inhibited (L-NAME, 100 µM) SLIGRL-induced, isolated EDHF responses are mediated by KCa3.1 alone (McNeish et al 2007). Under these conditions simvastatin had no effect on control EDHF hyperpolarization or relaxation (Control: 32.5 ± 3.6 mV and 76.1±4.7%; simvastatin 28.2 ± 3.8 mV and 75.6±3.1% n = 7). TRAM-34 only partially blocked EDHF responses (14.1±4.5 mV and 53.4±3.3% n = 7, P<0.05) and subsequent addition of apamin caused further block (7.7 ± 1.6 mVand36.5±8.5% n = 5, P<0.05). In the presence of L-NAME and simvastatin the isoprenoid gerenylgeranyl-pyrophosphate (GGPP, 1 µM) prevented the action of simvastatin as now TRAM-34 alone was required to block the EDHF response (Control: 22.5±5.5 mV and 76.6±2.3%; GGPP:23.9 ±3.2 mV and 76.9±2.9 %; GGPP+TRAM: 3.9±1.6 mV and 16.3±6.5%,P<0.05; n = 4). Statistics calculated using one-way ANOVA with Bonferroni’s post-test

Discussion: The data demonstrate a novel mechanism for regulation of KCa2.3 channels by statins. Stimulation of TP receptors involves ROCK signalling and inhibition of either TP (McNeish et al. 2007 and 2010) or ROCK restores KCa2.3 function to both EDH and isolated EDHF responses. Statins also restore this KCa2.3 component to EDH and EDHF but their actions are prevented by exogenous application of the isoprenoid GGPP, indicating that this effect is independent of cholesterol synthesis and likely to involve inhibition of Rho/ROCK mediated signalling. These findings are likely to have important implications for cardiovascular diseases associated with endothelial cell dysfunction such as stroke, hypertension and diabetes where statin treatment is beneficial. It may also help to explain some of the uncharacterised pleiotropic effects of statins in these diseases.

McNeish et al. (2010) Abstract submitted to winter BPS meeting.

McNeish et al. (2007) Br. J. Pharmacol. 151:441-449