Isolation and functional characterization of pericytes derived from hamster skeletal muscle

Aim: At the interface of tissue and capillaries, pericytes (PC) may generate electrical signals to be conducted along the skeletal muscle vascular network, but they are functionally not well characterized. We aimed to isolate and cultivate muscle PC allowing to analyse functional properties considered important for signal generation and conduction. Methods: Pericytes were enzymatically isolated from hamster thigh muscles and further selected during a 16-30 days' cultivation period. PC markers were studied by fluorescence activated cell scanning (FACS) and immunocytochemistry. Electrical properties of the cultured PC were investigated by patch clamp technique as well as the membrane potential sensitive dye DiBAC(4)(3). Results: The cultured cells showed typical PC morphology and were positive for NG2, alpha smooth muscle actin, PDGFR-beta and the gap junction protein Cx43. Expressions of at least one single or combinations of several markers were found in 80-90% of subpopulations. A subset of the patched cells expressed channel activities consistent with a Kv1.5 channel. In vivo presence of the channels was confirmed in sections of hamster thigh muscles. Interleukin-8, a myokine known to be released from exercising muscle, increased the expression but not the activity of this channel. Pharmacologic stimulation of the channel activity by flufenamic acid induced hyperpolarization of PC alone but not of endothelial cells [human umbilical vein endothelial cells (HUVEC)] alone. However, hyperpolarization was observed in HUVEC adjacent to PC when kept in co-culture. Conclusion: We established a culture method for PC from skeletal muscle. A first functional characterization revealed properties which potentially enable these cells to generate hyperpolarizing signals and to communicate them to endothelial cells.