The role of F-actin in hypo-osmotically induced cell volume change and calcium signaling in anulus fibrosus cells

Loading of the spine induces dynamic changes in the osmotic environment of the intervertebral disc (IVD) due to the exudation and recovery of tissue water. Cells from the anulus fibrosus (AF) respond to osmotic stress with altered biosynthesis through a pathway that may involve calcium (Ca2+) as a second messenger. We examined the hypothesis that AF cells respond to hypo-osmotic stress by swelling and initiating regulatory volume decrease (RVD). Further, the role of F-actin disruption and transient increases in intracellular calcium concentration ([Ca2+],) in volume adaptation were studied. In response to hypo-osmotic stress, AF cells swelled, disrupted F-actin, and exhibited [Ca2+], transients in proportion to the magnitude of the stress. The transient disruption of F-actin was dependent on the presence of extracellular Ca2+. After swelling, AF cells underwent RVD at all magnitudes of hypo-osmotic stress. The extent of RVD was diminished significantly by F-actin breakdown using cytochalasin D or by inhibition of swelling-induced F-actin disruption by removing extracellular Ca2+. Swelling-induced disruption of F-actin facilitated RVD, as evidenced by a more rapid volume recovery with increased F-actin breakdown. In conclusion, our findings suggest that the F-actin network plays an important role in the response of AF cells to osmotic stress.