The long-term objective of this project is to understand the nature of ion channel conductances in the basolateral membrane (BLM) of the renal proximal tubule (PT). ATP-sensitive potassium (KATP) channels underlie the dominant conductance (G) of the BLM. GK-ATP is a critical regulator of transport since the opening of BLM KATP channels maintains the driving force and provides a permeability required for continued epithelial transport. Therefore, in Aim I the biophysical characteristics of the ensemble conductance through BLM KATP channels will be determined, and other ion channels that contribute to GBLM will be identified. Since KATP channel activity is coupled to that of the Na+ pump, exploring the behavior of GK-ATP is crucial to understanding the regulation of transport in the PT. Therefore, in Aim 2 the regulation of the ensemble BLM KATP conductance by key cellular signaling pathways will be explored. This proposal focuses on defining the characteristics of the aggregate BLM KATP conductance by voltage clamping a large area of membrane (macropatch). The incentive to make macropatches on the BLM is clear: whole cell voltage clamping does not distinguish apical from BLM currents, and single channel recordings alone do not provide the signal to noise ratio, ease of analysis, or overall perspective that can be realized with macropatches. Although macropatches have been used in the study of cells with relatively smooth membranes (e.g., neurons), this approach has never been applied to epithelia due to technical obstacles. Therefore, the plan to exploit macropatches to explore GK ATP represents a novel, powerful approach that will allow reproducible and robust recordings of ensemble currents from the BLM that are unprecedented. The ultimate goal of these studies is to deepen our understanding of epithelial ion channels and how they are coupled to other membrane transport elements in coordinating overall cellular transport in the PT. Clinically, this work will provide insights relevant to hypertension, potassium balance, acute renal failure, and diabetes mellitus.
| Hurwitz, Craig G; Hu, Vivian Y; Segal, Alan S (2002) A mechanogated nonselective cation channel in proximal tubule that is ATP sensitive. Am J Physiol Renal Physiol 283:F93-F104 |
