Upregulation of apical NHE3 in renal OK cells overexpressing the rodent 1-subunit of the Na pump

Gomes, Pedro, and Patrı́cio Soares-da-Silva. Upregulation of apical NHE3 in renal OK cells overexpressing the rodent 1-subunit of the Na pump. Am J Physiol Regul Integr Comp Physiol 290: R1142–R1150, 2006. First published November 17, 2005; doi:10.1152/ajpregu.00102.2005.— Vectorial Na reabsorption across the proximal tubule is mediated by apical entry of Na , primarily via Na /H exchanger isoform 3 (NHE3), and basolateral extrusion via the Na pump (Na -K ATPase). We hypothesized that regulation of Na reabsorption should involve not only the activity of the basolateral Na -K ATPase, but also the apical NHE3, in a concerted manner. To generate a cell line that overexpresses Na -K -ATPase, opossum kidney (OK) cells were transfected with the rodent Na -K -ATPase 1-subunit (pCMV ouabain vector), and native cells were used as a control. The existence of distinct functional classes of Na -K -ATPase in wildtype and transfected cells was confirmed by the inhibition profile of Na -K -ATPase activity by ouabain. In contrast to wild-type cells, transfected cells exhibited two IC50 values for ouabain: the first value was similar to the IC50 of control cells, and the second value was 2 log units greater than the first, consistent with the presence of rat and opossum 1-isozymes. It is shown that transfection of OK cells with Na -K -ATPase increased Na -K -ATPase and NHE3 activities. This was associated with overexpression of the Na -K -ATPase 1-subunit and NHE3 in transfected OK cells. The abundance of the Na -K -ATPase 1-subunit was slightly lower in transfected OK cells. In conclusion, the increase in expression and function of Na -K -ATPase in cells transfected with the rodent Na pump 1-subunit cDNA is expected to stimulate apical Na influx into the cells, thereby accounting for the observed stimulation of the apical NHE3 activity.