Expression and function of sodium transporters in two opossum kidney cell clonal sublines

Abstract

The present study describes characteristic features of two clonal subpopulations of opossum kidney (OK) cells (OK(LC) and OK(HC)) that are functionally different but morphologically identical. The most impressive differences between OK(HC) and OK(LC) cells are the overexpression of Na+-K+-ATPase and type 3 Na+/H+ exchanger by the former, accompanied by an increased Na+-K+-ATPase activity (57.6 +/- 5.6 vs. 30.0 +/- 0.1 nmol P(i). mg protein(-1). min(-1)); the increased ability to translocate Na+ from the apical to the basolateral surface; and the increased Na+-dependent pH(i) recovery (0.254 +/- 0.016 vs. 0.094 +/- 0.011 pH units/s). Vmax values (in pH units/s) for Na+-dependent pHi recovery in OK(HC) cells (0.00521 +/- 0.0004) were twice (P < 0.05) those in OK(LC) (0.00202 +/- 0.0001), with similar Km values (in mM) for Na+ (OK(LC), 21.0 +/- 5.5; OK(HC), 14.0 +/- 5.6). In addition, we measured the activities of transporters (organic ions, alpha-methyl-D-glucoside, L-type amino acids, and Na+ and enzymes (adenylyl cyclase, aromatic L-amino acid decarboxylase, and catechol-O-methyltransferase). The cells were also characterized morphologically by optical and scanning electron microscopy and karyotyped. It is suggested that OK(LC) and OK(HC) cells constitute an interesting cell model for the study of renal epithelial physiology and pathophysiology, namely, hypertension.

Organ-Specific Overexpression of Renal LAT2 and Enhanced Tubular l -DOPA Uptake Precede the Onset of Hypertension

“…10,12,13 Another important aspect is concerned with the fact that Na ϩ is a powerful stimulus for the production of renal dopamine in humans and some laboratory animals, 40 and L-DOPA uptake in the human kidney is an Na ϩ -dependent and ouabain-sensitive process. 41 The finding that L-DOPA uptake in renal (LLC-PK 1 and OK cells) 17,27 and nonrenal (RBE4, DITNC1, and Neuro 2A) 42,43 cultured cells is largely promoted through the Na ϩ -insensitive, pH-dependent, and 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid-sensitive LAT2 transporter contrasts with the role of Na ϩ in the formation of renal dopamine. 40 In these cell types, most of the L-DOPA enters the cells in an Na ϩ -independent manner; only a minor component of L-DOPA uptake (Ϸ15%) was found to require extracellular Na ϩ .…”

“…This was particularly evident in OK HC cells, 17 which, like the renal tubular cells from hypertensive subjects, 44 are endowed with an enhanced ability to transport Na ϩ through the Na ϩ /H ϩ exchanger and Na ϩ ,K ϩ -ATPase. 43,44 …”

Na⁺/H⁺ Exchanger Activity and Dopamine D₁-Like Receptor Function in two Opossum Kidney Cell Clonal Sublines

“…The first evidence indicating differences between the two clones of OK cells, that are morphologically identical, was of the functional type and concerned their ability to take up L-DOPA, the precursor of dopamine, a local natriuretic hormone [9].The cells with the highest capacity to take up L-DOPA (OK HC cells) were those in that changes in transepithelial flux of Na + affected in a more important manner the uptake of L-DOPA [9].However, the most impressive difference between OK HC and OK LC cells was that the former over-expressed Na [10]. The characteristics of OK HC cells are of interest because some of these phenotypes have been described in renal proximal tubule cells from humans and rodents with genetic hypertension [11][12][13][14].…”

Giα3 protein-coupled dopamine D3receptor-mediated inhibition of renal NHE3 activity in SHR proximal tubular cells is a PLC-PKC-mediated event

“…Test compounds were added to the extracellular fluid during the acidification and Na ϩ -dependent pHi recovery periods. Intracellular pH measurements were performed in SHR cells cultured in polycarbonate filter supports or in 96-well plates (18). After loading the cells with 5 M BCECF-AM at 37°C for 30 min, test compounds were added to the extracellular fluid 25 min before starting the sodium-dependent pH i recovery period.…”