Mechanoelectrical transduction in chondrocytes.

Cartilage tissue lines the joints of mammals, helping to lubricate joint movement and distribute mechanical loads. This tissue is comprised of isolated cells known as chondrocytes which are embedded in an extracellular matrix. Chondrocytes produce and maintain the cartilage by sensing and responding to changing mechanical loads. Mechanosensitive ion channels have been implicated in chondrocyte mechanotransduction and recent studies have shown that both PIEZO1 and TRPV4 can be activated by mechanical stimuli in these cells. The 2 channels mediate separate but overlapping mechanoelectrical transduction pathways, PIEZO1 in response to stretch and substrate deflections and TRPV4 in response to substrate deflections alone. These distinct pathways of mechanoelectrical transduction suggest a mechanism by which chondrocytes can distinguish between different stimuli that arise in their complex mechanical environment.

Acrosome reaction and Ca²âº imaging in single human spermatozoa: new regulatory roles of [Ca²âº]i.

The spermatozoa acrosome reaction (AR) is essential for mammalian fertilization. Few methods allow visualization of AR in real time together with Ca²âº imaging. Here, we show that FM4-64, a fluorescent dye used to follow exocytosis, reliably reports AR progression induced by ionomycin and progesterone in human spermatozoa. FM4-64 clearly delimits the spermatozoa contour and reports morphological cell changes before, during, and after AR. This strategy unveiled the formation of moving tubular appendages, emerging from acrosome-reacted spermatozoa, which was confirmed by scanning electron microscopy. Alternate wavelength illumination allowed concomitant imaging of FM4-64 and Fluo-4, a Ca²âº indicator. These AR and intracellular Ca²âº ([Ca²âº]i) recordings revealed that the presence of [Ca²âº]i oscillations, both spontaneous and progesterone induced, prevents AR in human spermatozoa. Notably, the progesterone-induced AR is preceded by a second [Ca²âº]i peak and ~40% of reacting spermatozoa also manifest a slow [Ca²âº]i rise ~2 min before AR. Our findings uncover new AR features related to [Ca²âº]i.

Direct measurement of TRPV4 and PIEZO1 activity reveals multiple mechanotransduction pathways in chondrocytes.

The joints of mammals are lined with cartilage, comprised of individual chondrocytes embedded in a specialized extracellular matrix. Chondrocytes experience a complex mechanical environment and respond to changing mechanical loads in order to maintain cartilage homeostasis. It has been proposed that mechanically gated ion channels are of functional importance in chondrocyte mechanotransduction; however, direct evidence of mechanical current activation in these cells has been lacking. We have used high-speed pressure clamp and elastomeric pillar arrays to apply distinct mechanical stimuli to primary murine chondrocytes, stretch of the membrane and deflection of cell-substrate contacts points, respectively. Both TRPV4 and PIEZO1 channels contribute to currents activated by stimuli applied at cell-substrate contacts but only PIEZO1 mediates stretch-activated currents. These data demonstrate that there are separate, but overlapping, mechanoelectrical transduction pathways in chondrocytes.