MEASUREMENTS OF GROWTH CONE ADHESION TO CULTURE SURFACES BY MICROMANIPULATION

Neurons were grown on plastic surfaces that were untreated, or treated with polylysine, laminin, or L1 and their growth cones were detached from their culture surface by applying known forces with calibrated gl ass needles. This detachment force was taken as a measure of the force of adhesion of the growth cone. We find that on all surfaces, lamelli podial growth cones require significantly greater detachment force tha n filopodial growth cones, but this difference is, in general, due to the greater area of lamellipodial growth cones compared to filopodial growth cones. That is, the stress (force/unit area) required for detac hment was similar for growth cones of lamellipodial and filopodial mor phology on all surfaces, with the exception of lamellipodial growth co nes on L1-treated surfaces, which had a significantly lower stress of detachment than on other surfaces. Surprisingly, the forces required f or detachment (760-3,340 mu dynes) were three to 15 times greater than the typical resting axonal tension, the force exerted by advancing gr owth cones, or the forces of retraction previously measured by essenti ally the same method. Nor did we observe significant differences in de tachment force among growth cones of similar morphology on different c ulture surfaces, with the exception of lamellipodial growth cones on L 1-treated surfaces. These data argue against the differential adhesion mechanism for growth cone guidance preferences in culture. Our microm anipulations revealed that the most mechanically resistant regions of growth cone attachment were confined to quite small regions typically located at the ends of filopodia and lamellipodia. Detached growth con es remained connected to the substratum at these regions by highly ela stic retraction fibers. The closeness of contact of growth cones to th e substratum as revealed by interference reflection microscopy (IRM) d id not correlate with our mechanical measurements of adhesion, suggest ing that IRM cannot be used as a reliable estimator of growth cone adh esion.