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 in intracellular calcium concentration ([Ca²⁺](i)) evoked by elevating external [Ca²⁺] from 10 nM to 1 mM (Δ[Ca²⁺]). Under control conditions, Δ[Ca²⁺] averaged 6 nM and was unaffected by elevating bath [K⁺]. After treatment with 1 μM thapsigargin to deplete the intracellular Ca²⁺ store, the change in [Ca²⁺](i) (Δ[Ca²⁺](th)) averaged 147 ± 16 nM. In thapsigargin-treated MC Studied under depolarizing conditions (75 mM bath K⁺), Δ[Ca²⁺](th) was 45 ± 7 nM. The Δ[Ca²⁺](th) response of thapsigargin-treated cells was inhibited by La³⁺ (IC₅₀ = 335 nM) but was unaffected by 5 μM Cd²⁺. In patch clamp studies, inward currents were observed in cell-attached 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 μM La³⁺, insensitive to 5 μM Cd²⁺, and voltage independent, with an average channel activity (NP₀) 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 through SOC, as well as a small- conductance divalent-selective channel displaying biophysical properties consistent with SOC. Based on estimates of whole cell Ca²⁺ influx derived from our data, we conclude that SOC with low single-channel conductance must be highly abundant in MC to allow significant capacitative Ca²⁺ entry in response to depletion of the intracellular store.