GUIDANCE OF CNS GROWTH CONES BY SUBSTRATUM GROOVES AND RIDGES - EFFECTS OF INHIBITORS OF THE CYTOSKELETON, CALCIUM CHANNELS AND SIGNAL-TRANSDUCTION PATHWAYS

We exploited our observation that embryonic Xenopus spinal neurites al ign parallel to grooves in a quartz surface and that embryonic rat hip pocampal neurites align perpendicular to shallow, narrow grooves (see companion paper: A, M. Rajnicek, S. Britland and C. D. McCaig, 1997) ( J. Cell Sci. 110, 2905-2913) to investigate the mechanism of growth co ne contact guidance, Substratum topography affected the pattern of gro wth cone filopodia and microtubules but parallel orientation of Xenopu s neurites and perpendicular orientation of hippocampal neurites were unperturbed by cytochalasin B, which virtually eliminated filopodia, H ippocampal growth cone orientation and turning in response to grooves was unaffected by disruption of microtubules using taxol or nocodazole . Gross cytoskeletal reorganization on grooved substrata was therefore not required for growth cone steering. Inhibitors were used to identi fy the signal transduction pathway for perpendicular alignment of hipp ocampal neurites, Alignment persisted in the presence of gadolinium ch loride, a blocker of stretch-activated calcium channels, the G protein inhibitor pertussis toxin, the protein tyrosine kinase inhibitor geni stein, the protein kinase A and G inhibitor HA1004, the protein kinase A inhibitor KT5720 and the protein kinase G inhibitor KT5823. Low con centrations of the protein kinase C inhibitors staurosporine, bisindol ylmaleimide or H-7 did not affect perpendicular orientation but higher concentrations inhibited it, The calcium channel blockers flunarizine , nifedipine and diltiazem also inhibited perpendicular orientation, I nflux of calcium and protein kinase C activity therefore appear to be involved in perpendicular contact guidance.