9513958 McDonough Potassium (K+) is the main salt found in intracellular fluid (ICF), while extracellular fluid (ECF) potassium is low, so there is a gradient of potassium across the cell membrane which is necessary for nerve and muscle excitability. Vertebrate animals must regulate extracellular fluid potassium concentration (ECF K+ ) within a narrow range. If K+ loss exceeds input over a period of days, ECF K+ falls, which is known as hypokalemia. The resulting increase in the K+ gradient across the cell membrane, if not corrected, can disrupt critical functions such as cardiac contractility (which can be fatal). During potassium deprivation, the skeletal muscle adapts by providing its K+ to the ECF to blunt and delay the fall in ECF K+. This important regulatory process is only poorly understood. In contrast, non-muscle cell types are adapted to maintain, not lose, intracellular K+ during K+ deprivation. The central aim of this proposal is to identify the specific muscle types that supply K+ to the ECF in hypokalemia and the underlying cellular mechanisms responsible for the shift and for the restoration response when K+ becomes available. Defining the target muscles and transporters are critical first steps toward understanding why a subset of muscles is specialized to lose K+ while others are spared in order to perform, different critical muscle functions. The loss of ICF K+ is accompanied by a decrease in number of sodium pumps, the molecules in the membrane that actively pump K+ into the cell from the ECF. This supports the hypothesis that the K+ loss is due to a decrease in active K+ influx rather than an increase in K+ exit from the cell. Sodium pumps, Na,K-ATPase, are composed of ( and ( subunits and muscle expresses (l, (2, (1, (2 subunit isoforms, thus, up to four possible (( combinations. The PI's laboratory has established that the isoforms are expressed in a muscle specific pattern. The PI has further established that abundance of (2 and (2 subunits are depressed in a muscle specific pattern. The PI aims to accomplish the following: 1) Test the hypothesis that the decrease in number of specific sodium pump isoforms is the underlying cellular mechanism causing shifts in K+ from a subset of muscles to ECF in hypokalemia. The PI will determine time course and muscle specificity of change in: ICF K+ , K+ influx, K+ efflux, and sodium pump subunit isoform abundance during low K+ diet and after restoring K+ in a panel of five muscles including white gastrocnemius where there is a 85% decrease in (2(2, to diaphragm where there is no change in (2(2 in hypokalemia. 2) Test the hypothesis that muscle sodium pump activity is regulated at multiple levels, both long term regulation of RNA and protein pool size, and acute regulation of existing pumps, during hypokalemia and K+ restoration. During K+ depiction and repletion determine the time course of change in ( and ( isoforms' mRNA vs. protein, and determine whether there is also acute regulation of Na,K-ATPase transport activity before there are changes in abundance of pumps. Significance: this fundamental, specialized, and poorly understood response of skeletal muscle to organism K+ deprivation is critical for survival of all vertebrates, including humans, during periods of fasting or famine, or when K+ loss is high (mineralocorticoids, prolonged exercise in the heat, diuretics). Accomplishing these aims will identify the location of the K+ reservoir and the cellular mechanisms responsible for tapping and repleting this reservoir. This information contributes to an understanding how a subset of muscles have specialized to lose K+ to buffer ECF K+ , thus sparing other muscles and organs to perform critical functions, and to development of assays for the signal(s) that mediate the response. All steps are requisite for defining the homeostatic loops responsible for adapting to altered K+ status. ***
