Abstract
Adult guinea-pig ventricular myocytes (aGPVMs) exhibit enduring action-potential plateaus similar to those in humans, making this system advantageous for investigating human phase 2 phenomena. Long-term cultures of aGPVMs (Canadian J Physiol Pharmacol7:740) further offer the promise of genetic manipulation by somatic gene transfer, but have been infrequently exploited. Here, we confirm that aGPVM cultures form monolayers featuring organized myofibrils and robust contraction. Beyond this, we now demonstrate that point pacing of di-4-ANEPPS stained monolayers enable optical measurements of action potentials. At 2-Hz pacing, action potentials indeed lasted ∼250 ms, but displayed attenuated plateau phases (a, gray trace). To normalize phase 2, and to demonstrate genetic manipulability, we virally expressed Ca2+ channel CaVβ subunits that enhance L-type channels in heterologous systems. CaVβ expression in fact restored a full-bore plateau phase, as shown by the black trace in a. To confirm a role of Ca2+ current, we obtained single-channel recordings of native L-type channels. CaVβ subunits strikingly increased open probability, while decreasing inactivation (b, control; c, CaVβ expression), thus explaining the normalization of action-potential morphology. Overall, long-term cultured aGPVMs offer a powerful model system for electrophysiological exploration.
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