Visualization and Quantitation of Neurotrophin mRNAs

During development of the vertebrate nervous system, neurons are produced in excess and, at a restricted time period, a significant portion degenerate, a phenomenon referred to as naturally occurring cell death (Hamburger, 1975). The neurons that survive this phase are those that form functional connections with their targets, competing successfully for a limited supply of a target-derived trophic factor (Barde, 1989). The concept of neurotrophic interactions in the nervous system has emerged largely from studies on nerve growth factor (NGF), a wellcharacterized trophic factor that in the PNS supports neural crest-derived sensory neurons and sympathetic neurons (LeviMontalcini, 1987). Apart from NGF, indirect evidence for the existence of many other neurotrophic factors has been obtained from numerous in vitro experiments using various tissue extracts (Dohrman et al., 1986; Tomozawa and Appel, 1986) or cocultures (Prochiantz et al., 1979) to demonstrate support of neuronal development and survival. Thus, it was considered likely that other neurotrophic factors exist and the purification of brainderived neurotrophic factors (BDNF) (Barde et al., 1982) demonstrated this to be true. The subsequent isolation of the gene for BDNF (Leibrock et al., 1989) led to the striking observation that the deduced primary structure of BDNF showed strong similarity to NGF, suggesting the possible existence of a gene family of related neurotrophic factors. The homologous sequences were used by several groups to isolate a third member of this neurotrophic factor family, neurotrophin-3 (NT-3) (Emfors et al., 1990a; Hohn et al., 1990; Kaisho et al., 1990; Maisonpierre et al,, 1990a; Rosenthal et al., 1990). The gene for a fourth member (neurotrophin-4; NT-4) of this family was first isolated and characterized from Xenopus laevis and Vipera lebetina (Hallb��k et al. 1991). Recently, a mammalian homologue of NT-4 has been molecularly cloned from rat and human (Ip et al., 1992). The same sequence was also recently isolated by Berkemeier et al. (1991) who named it neurotrophin-5 (NT-5). However, the fact that this sequence is substantially more related to Xenapus NT-4