No abstract

We have investigated the co-involvement of endogenous NGF and impulses in the collateral sprouting of cutaneous sensory nerves in adult rats, specifically the A delta-axons involved in mechanonociception and the C-fibers that mediate heat nociception. Their collateral sprouting was measured by the progressive expansion, respectively, of the behaviorally defined "pinch" and "heat" fields into surrounding denervated skin (the light-touch A alpha-fibers do not sprout in adult mammals). The expansions of such "isolated" fields were totally prevented in animals injected daily with anti-NGF serum, but developed normally after treatment was discontinued. Light microscopic and EM examination of the skin confirmed that the effect of the anti-NGF treatment was attributable to its prevention of collateral sprouting. Initiation of treatment would also rapidly halt sprouting already in progress. Finally, intradermal injections of purified NGF protein would not only increase the rate of nociceptive fiber sprouting, but also evoke sprouting de novo within normally innervated skin (again, A alpha-axons were unaffected). We conclude that the collateral sprouting of intact nociceptive nerves following partial denervation of skin is entirely dependent on endogenous NGF. The observed latency of this sprouting was 10-12 d; we estimate, however, that at least 2 d of field expansion is required for its reliable detection. Thus, about 8-10 d are required for NGF levels in the skin to rise to effective levels, and for the neurons to respond and initiate sprouting. From indirect findings, the NGF component of this sprouting latency appears to be about 2 d. In accord with earlier findings, the remaining "initiation time" was reduced by 5-6 d if the neurons were briefly excited, even 2 d prior to the isolation of their fields. Unexpectedly, this phenomenon of "precocious sprouting" requires that endogenous NGF be available; the sprouting latency reverted to normal values when the conditioning impulses were evoked during a 2 d anti-NGF "umbrella." In contrast to the impulse-sensitive neuronal mechanisms involved in the initiation of sprouting, those underlying the sprouting rate were unaffected by nerve activity and were entirely dependent on the level of endogenous NGF. We suggest that interactions like that revealed in these studies between a sprouting agent and impulses that seem to prime the neuron's response to it contribute to plasticity within the nervous system.

Nerve growth factor (NGF) promotes neuronal survival and differentiation and stimulates neurite outgrowth. NGF is synthesized as a precursor, proNGF, which undergoes posttranslational processing to generate mature b-NGF. It has been assumed that, in vivo, NGF is largely processed into the mature form and that mature NGF accounts for the biological activity. However, we recently showed that proNGF is abundant in CNS tissues whereas mature NGF is undetectable, suggesting that proNGF has biological functions beyond its role as a precursor. To determine whether proNGF exhibits biological activity, we mutagenized the precursor-processing site and expressed unprocessed, cleavage-resistant proNGF protein in insect cells. Survival and neurite outgrowth assays on murine superior cervical ganglion neurons and PC12 cells indicated that proNGF exhibits neurotrophic activity similar to mature 2.5S NGF, but is approximately fivefold less active. ProNGF binds to the high-affinity receptor, TrkA, as determined by cross-linking to PC12 cells, and is also slightly less active than mature NGF in promoting phosphorylation of TrkA and its downstream signaling effectors, Erk1/2, in PC12 and NIH3T3-TrkA cells. These data, coupled with our previous report that proNGF is the major form of NGF in the CNS, suggest that proNGF could be responsible for much of the biological activity normally attributed to mature NGF in vivo.

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