Substrate‐bound factors stimulate engorgement of growth cone lamellipodia during neurite elongation

Abstract

The surfaces on which neurons grow greatly affect neurite elongation, but it is unclear how substrates influence the events within the growth cone that bring about elongation. Neurite elongation by Aplysia californica neurons in culture occurs through a series of transformations of the structures of the growth cone (Goldberg and Burmeister, J. Cell Biol., 103:1921-1931, 1986). The growth cone produces actin-rich protrusions, veils, and lamellipodia, which can then mature into the central body of the growth cone through the net advance of microtubules and membranous organelles from contiguous central regions, a process called "engorgement." Aplysia neurons form growth cones on poly-l-lysine-treated substrates, but their rate of neurite elongation is greatly enhanced on substrates additionally exposed to Aplysia hemolymph. The acute application of hemolymph to slowly growing neurites brings about a rapid acceleration of neurite elongation and engorgement. The enhancement of engorgement was effected with material eluted from hemolymph-treated substrates and was not seen when hemolymph was added to neurons cultured on hemolymph-treated substrates inactivated by exposure to UV radiation. Thus, we conclude that the rapid acceleration of engorgement caused by hemolymph is, in large part, a substrate-mediated effect. We propose that extracellular substrate molecules can modulate the rate of neurite growth through the regulation of the engorgement of lamellipodia. The microtubule disrupters colcemid and nocodazole inhibit the advance of vesicular elements into the lamellipodia following hemolymph treatment, but taxol, which promotes the polymerization and stabilization of microtubules, does not itself enhance engorgement. The microfilament disrupter cytochalasin B, however, stimulates engorgement. Our results suggest that regulating the resistance of the peripheral actin meshwork to penetration by microtubules and vesicles may be a mechanism by which substrate-attached molecules regulate neurite advance.

Arterial cells and CNS sheath cells from Aplysia californica produce factors that enhance neurite outgrowth in co-cultured neurons

“…5B, C, 6A). Culture medium containing 50% hemolymph strongly promotes growth by Aplysia neurons (Burmeister et al 1991;Goldberg and Schacher 1998;Schacher 1988;Schacher and Proshansky 1983). Culture medium consisting of SL-15 alone was sufficient to sustain minimal growth; however, no significant change in the percentage of neurons with growth occurred in this condition throughout the 48-h culture period (logistic regression analysis, P<0.01).…”

“…One possibility is that the diffusible factor produced in arterial cell cultures is a soluble form of acetylcholinesterase, which is present in Aplysia hemolymph and enhances neurite growth from isolated Aplysia neurons (Srivatsan and Peretz 1997). It is also possible to establish a substrate-bound form of growth factor(s) from Aplysia hemolymph (Burmeister et al 1991), and it may be that the soluble form of acetylcholinesterase binds to coated glass coverslips and can promote neurite outgrowth. Given their large size and abundance, the source of the secreted soluble factor(s), in our cultures as well as in hemolymph, may be the arterial myocytes.…”

Endogenous neurotrophic factors enhance neurite growth by bag cell neurons of Aplysia

“…Also, Burmeister et al (1991) show that Aplysia hemolymph can be used to establish a substrate on glass coverslips capable of promoting extensive neurite growth by isolated Aplysia neurons. In separate studies, conditioned medium derived from the Aplysia CNS elicits extensive neurite outgrowth from isolated Aplysia neurons (Wong et al, 1983;Fejtl and Carpenter, 1995).…”

“…Mollusk-derived growth factor is expressed in the reproductive tract and CNS of Aplysia during juvenile development, and has been implicated in CNS repair after injury (Akalal and Nagle, 2001). Also, a soluble form of acetylcholine esterase is found in Aplysia hemolymph, and it enhances neurite outgrowth from Aplysia neurons (Burmeister et al, 1991;Svrivatsan and Peretz, 1997;Srivatsan, 1999). No endogenous receptors for the aforementioned growth factors have been identified (McKay et al, 1999).…”

“…Schacher (1981) reported that support cells of Aplysia play a role in the growth and differentiation of neurons during development of the abdominal ganglion in Aplysia. Also, hemolymph of Aplysia contains factors that enhance neurite outgrowth (Wong et al, 1983;Burmeister et al, 1991;Srivatsan and Peretz, 1997).…”