Factors Affecting Membrane Permeability and Ionic Homeostasis in the Cold-Submerged Frog

Frogs (Rana temporaria) were submerged at 3 °C in either normoxic or hypoxic water for up to 16 weeks, and denied air access, to mimic the conditions of an ice-covered pond during the winter. The activity of the skeletal muscle Na⁺/K⁺ pump over the first 2 months of hibernation, measured by ouabain-inhibitable ²²Na⁺ efflux, was reduced by 30 % during normoxia and by up to 50 % during hypoxia. The reduction in Na⁺/K⁺ pump activity was accompanied by reductions in passive ²²Na⁺ influx and ⁸⁶Rb⁺ efflux (effectively K⁺ efflux) across the sarcolemma. This may be due to a decreased Na⁺ permeability of the sarcolemma and a 75 % reduction in K⁺ leak mediated by ATP-sensitive K⁺ channels (‘K_ATP’ channels). The lowered rates of ²²Na⁺ and ⁸⁶Rb⁺ flux are coincident with lowered transmembrane ion gradients for [Na⁺] and [K⁺], which may also lower Na⁺/K⁺ pump activity. The dilution of extracellular [Na⁺] and intracellular [K⁺] may be partially explained by increased water retention by the whole animal, although measurements of skeletal muscle fluid compartments using ³H-labelled inulin suggested that the reduced ion gradients represented a new steady state for skeletal muscle. Conversely, intracellular ion homeostasis within ventricular muscle was maintained at pre-submergence levels, despite a significant increase in tissue water content, with the exception of the hypoxic frogs following 4 months of submergence. Both ventricular muscles and skeletal muscles maintained resting membrane potential at pre-submergence levels throughout the entire period of hibernation. The ability of the skeletal muscle to maintain its resting membrane potential, coincident with decreased Na⁺/K⁺ pump activity and lowered membrane permeability, provided evidence of functional channel arrest as an energy-sparing strategy during hibernation in the cold-submerged frog.