Enhancement of PC12 axonal extension via hybrid electromagnetic and mechanical stimulation

In this study, a hybrid electromagnetic and mechanical stimulation system that can apply an alternative current magnetic field (ACMF) and tensile strain fields on PC12 cells was developed to enhance nerve axonal extension. For the ACMF stimulation system, we used a frame to facilitate uniform ACMF application and in situ microscopic observation. We optimized the design of the frame based on analytical results. We verified that the developed ACMF stimulation system can generate a uniform magnetic field. Further, we designed a uniaxial stretch stimulation system. The cell culture area of the stretch stimulation system was made of a nonmagnetic material. The strain in the stretch stimulation region was confirmed to be uniform, with acceptably small deviations. Next, the effectiveness of axonal extension enhancement was validated by adopting two stimulation methods, ACMF and stretch, separately or in combination, hereafter referred to as ACMF, stretch, and hybrid conditions. PC12 cells seeded on the silicone sheets were cultured for 96 h under the three stimulation conditions. The enhancement rate of the hybrid condition was higher than the enhancement rates of ACMF stimulation and stretch stimulation. The effects of stretch stimulation on axonal extension appeared immediately after beginning the stimulation, while the effects of ACMF stimulation took longer to appear. These results revealed that there are different mechanisms of cell environment stimulation of the axonal extension of PC12 cells and that hybrid stimulation is the most effective stimulation method of those studied.