Store-operated Ca2+ channels in human glomerular mesangial cells

Experiments were performed to identify the biophysical properties of store- operated Ca2+ channels (SOC) in cultured human glomerular mesangial cells ( MC). A fluorometric technique (fura 2) was utilized to monitor the change i n intracellular calcium concentration ([Ca2+](i)) evoked by elevating exter nal [Ca2+] from 10 nM to 1 mM (Delta[Ca2+]). Under control conditions, Delt a[Ca2+] averaged 6 nM and was unaffected by elevating bath [K+]. After trea tment with 1 mu M thapsigargin to deplete the intracellular Ca2+ store, the change in [Ca2+](i) (Delta[Ca2+](th)) averaged 147 +/- 16 nM. In thapsigar gin-treated MC studied under depolarizing conditions (75 mM bath K+), Delta [Ca2+](th) was 45 +/- 7 nM. The Delta[Ca2+](th) response of thapsigargin-tr eated cells was inhibited by La3+ (IC50 = 335 nM) but was unaffected by 5 m u M Cd2+. In patch clamp studies, inward currents were observed in cell-att ached patches with either 90 mM Ba2+ or Ca2+ in the pipette and 140 mM KCl in the bathing solution. The single-channel conductance was 2.1 pS with Ba2 + and 0.7 pS with Ca2+. The estimated selectivities were Ca2+ > Ba2+ >> K+. These channels were sensitive to 2 mu M La3+, insensitive to 5 mu M Cd2+, and voltage independent, with an average channel activity (NPo) of 1.02 at command potential (-V-p) ranging from 0 to -80 mV. In summary, MC exhibited an electrogenic Ca2+ influx pathway that is suggestive of Ca2+ entry throu gh SOC, as well as a small-conductance divalent-selective channel displayin g biophysical properties consistent with SOC. Based on estimates of whole c ell Ca2+ influx derived from our data, we conclude that SOC with low single -channel conductance must be highly abundant in MC to allow significant cap acitative Ca2+ entry in response to depletion of the intracellular store.