Fluid shear stress induces calcium transients in osteoblasts through depolarization of osteoblastic membrane.

Intracellular calcium transient ([Ca(2+)]i transient) induced by fluid shear stress (FSS) plays an important role in osteoblastic mechanotransduction. Changes of membrane potential usually affect [Ca(2+)]i level. Here, we sought to determine whether there was a relationship between membrane potential and FSS-induced [Ca(2+)]i transient in osteoblasts. Fluorescent dyes DiBAC4(3) and fura-2AM were respectively used to detect membrane potential and [Ca(2+)]i. Our results showed that FSS firstly induced depolarization of membrane potential and then a transient rising of [Ca(2+)]i in osteoblasts. There was a same threshold for FSS to induce depolarization of membrane potential and [Ca(2+)]i transients. Replacing extracellular Na(+) with tetraethylammonium or blocking stretch-activated channels (SACs) with gadolinium both effectively inhibited FSS-induced membrane depolarization and [Ca(2+)]i transients. However, voltage-activated K(+) channel inhibitor, 4-Aminopyridine, did not affect these responses. Removing extracellular Ca(2+) or blocking of L-type voltage-sensitive Ca(2+) channels (L-VSCCs) with nifedipine inhibited FSS-induced [Ca(2+)]i transients in osteoblasts too. Quantifying membrane potential with patch clamp showed that the resting potential of osteoblasts was -43.3mV and the depolarization induced by FSS was about 44mV. Voltage clamp indicated that this depolarization was enough to activated L-VSCCs in osteoblasts. These results suggested a time line of Ca(2+) mobilization wherein FSS activated SACs to promote Na(+) entry to depolarize membrane that, in turn, activated L-VSCCs and Ca(2+) influx though L-VSCCs switched on [Ca(2+)]i response in osteoblasts.