Interrogating the Molecular Clutch in Neuronal Growth Cones: Measuring Traction Forces, F-actin Retrograde Flow, and Point Contact Demographics

Growth cone-dependent outgrowth of neuronal processes is essential for the development, plasticity, and regenerative capacity of the nervous system. This process involves the attachment of the growth coneGrowth cones to the substrate and the cyclical engagement/disengagement of the molecular clutchMolecular clutches at the sites of adhesive contact. In this chapter, we describe protocols for traction force microscopyTraction force microscopy (TFM), measurement of F-actin retrograde flowF-actin retrograde flow velocities, and the assessment of adhesive point contacts by immunofluorescenceImmunofluorescence. These complementary techniques collectively facilitate investigations into the regulation of the molecular clutchMolecular clutches in neuronal growth conesGrowth cones.