Hyperosmotically induced volume change and calcium signaling in intervertebral disk cells: The role of the actin cytoskeleton

Loading of the spine alters the osmotic environment in the intervertebral disk (IVD) as interstitial water is expressed from the tissue. Cells from the three zones of the IVD, the anulus fibrosus (AF), transition zone (TZ), and nucleus pulposus (NP), respond to osmotic stress with altered biosynthesis through a pathway that may involve calcium (Ca²⁺) as a second messenger. We examined the hypothesis that IVD cells respond to hyperosmotic stress by increasing the concentration of intracellular calcium ([Ca²⁺]_i) through a mechanism involving F-actin. In response to hyperosmotic stress, control cells from all zones decreased in volume and cells from the AF and TZ exhibited [Ca²⁺]_i transients, while cells from the NP did not. Extracellular Ca²⁺ was necessary to initiate [Ca²⁺]_i transients. Stabilization of F-actin with phalloidin prevented the Ca²⁺ response in AF and TZ cells and decreased the rate of volume change in cells from all zones, coupled with an increase in the elastic moduli and apparent viscosity. Conversely, actin breakdown with cytochalasin D facilitated Ca²⁺ signaling while decreasing the elastic moduli and apparent viscosity for NP cells. These results suggest that hyperosmotic stress induces volume change in IVD cells and may initiate [Ca²⁺]_i transients through an actin-dependent mechanism.