Andrej G. Kamkin

Vascular smooth muscle voltage-gated potassium (Kv) channels have been proposed to contribute to myogenic autoregulation. Surprisingly, in initial experiments, we observed that the Kv2 channel inhibitor stromatoxin induced vasomotion without affecting myogenic tone. Thus, we tested the hypothesis that Kv2 channels contribute to myogenic autoregulation by fine-tuning the myogenic response. Expression of Kv2 channel mRNA was determined using real-time PCR and ‘multiplex’ single-cell RT-PCR. Potassium currents were measured using the patch-clamp technique. Contractile responses of intact arteries were studied using isobaric myography. Expression of Kv2.1 but not Kv2.2 channels was detected in intact rat superior cerebellar arteries and in single smooth muscle cells. Stromatoxin, a high-affinity inhibitor of Kv2 channels, reduced smooth muscle Kv currents by 61% at saturating concentrations (EC₅₀ 36 nmol/L). Further, stromatoxin (10–100 nmol/L) induced pronounced vasomotion in 48% of the vessels studied. In vessels not exhibiting vasomotion, stromatoxin did not affect myogenic reactivity. Notably, in vessels exhibiting stromatoxin-induced vasomotion, pressure increases evoked two effects: First, they facilitated the occurrence of random vasodilations and/or vasoconstrictions, disturbing the myogenic response (24% of the vessels). Second, they modified the vasomotion by decreasing its amplitude and increasing its frequency, thereby destabilizing myogenic tone (76% of the vessels). Our study demonstrates that (i) Kv2.1 channels are the predominantly expressed Kv channels in smooth muscle cells of rat superior cerebellar arteries, and (ii) Kv2.1 channels provide a novel type of negative feedback mechanism in myogenic autoregulation by preventing vasomotion and thereby safeguarding the myogenic response.

We studied the effect of anti-inflammatory ininterleukin-13 (IL-13; 50 ng/ml) on bioelectrical activity of rat atrial cardiomyocytes under control conditions and on the background of stretch by means of microelectrode technique. IL-13 did not lead to alterations of the resting membrane potential and action potential amplitude during 35 minutes. However APD25, APD50, APD90 in 50% of cases significantly decreased, while in other 50% of cases it increased. In case when IL-13 decreased APD cellular stretch by 1.7 mN caused an increase in APD frequency by 120% and caused decrease in APD25, APD50, APD90, which happened on the background of modest cellular depolarization in the range of 5 mV. When IL-13 increased APD, tissue stretching just by 0.8 mN caused depression of the frequency by 10% and increase in APD25, APD50, APD90. This happened on the background of small cellular depolarization.

The role of NO in the regulation of currents passing through ion channels activated by cell stretching (mechanically gated channels, MGC), particularly through cation-selective K⁺-channels TRPC6, TREK1 (K_2P2.1), and TREK2 (K_2P10.1), was studied on isolated mouse, rat, and guinea pig cardiomyocytes using whole-cell patch-clamp technique. In non-deformed cells, binding of endogenous NO with PTIO (2-(4-carboxyphenyl)-4,4,5,5-tetramethyl-imidazoline- 1-1-oxy-3-oxide) irreversibly shifted the diastolic membrane potential towards negative values, modulates Kir-channels by reducing I_K1, and blocks MGC. Perfusion of stretched cells with PTIO solution completely blocked MG-currents. NO-synthase inhibitors L-NAME and L-NMMA completely blocked MGC. Stretching of cardiomyocytes isolated from wild type mice and from NOS1^–/–- and NOS2^–/–- knockout mice led to the appearance in MG-currents typical for the specified magnitude of stretching, while stretching of cardiomyocytes from NOS3–/–- knockout mice did not produce in MG-current. These findings suggest that NO plays a role in the regulation of MGC activity and that endothelial NO-synthase predominates as NO source in cardiomyocyte response to stretching.

Whole-cell ionic currents through mechanically gated chnnels (MGC) were recorded in isolated cardiomyocytes under voltage clamp conditions. In unstrained cells, NO donors SNAP and DEA-NO activated MGC and induced MG-like currents. In contrast, in stretched cells with activated MGC, these NO-donors inactivated and inhibited MGC.