Developmental regulation of neuronal K+ channels by target-derived TGF beta in vivo and in vitro.

The functional expression of Ca2+-activated K+ channels (KCa) in developing chick ciliary ganglion (CG) neurons requires interactions with target tissues and preganglionic innervation. Here, we show that the stimulatory effects of target tissues are mediated by an isoform of TGFbeta. Exposure of cultured CG neurons to TGFbeta1, but not TGFbeta2 or TGFbeta3, caused robust stimulation of KCa. The KCa stimulatory effects of target tissue extracts were blocked by a neutralizing pan-TGFbeta antiserum but not by specific TGFbeta2 or TGFbeta3 antisera. Intraocular injection of TGFbeta1 caused robust stimulation of KCa, whereas intraocular injection of pan-TGFbeta antiserum inhibited expression of KCa in CG neurons developing in vivo. The effects of TGFbeta1 were potentiated by beta-neuregulin-1, a differentiation factor expressed in preganglionic neurons.

Neuregulins stimulate the functional expression of Ca2+-activated K+ channels in developing chicken parasympathetic neurons.

The developmental expression of macroscopic Ca2+-activated K+ currents (IK[Ca]) in chicken ciliary ganglion (CG) neurons is dependent in part on trophic factors released from preganglionic nerve terminals. Neuregulins are expressed in the preganglionic neurons that innervate the chicken CG and are therefore plausible candidates for this activity. Application of 1 nM beta1-neuregulin peptide for 12 hr evokes a large (7- to 10-fold) increase in IK[Ca] in embryonic day 9 CG neurons, even in the presence of a translational inhibitor. A similar posttranslational effect is produced by high concentrations (10 nM) of epidermal growth factor and type alpha transforming growth factor but not by 10 nM alpha2-neuregulin peptide or by neurotrophins at 40 ng.ml-1. beta1-neuregulin treatment for 12 hr also confers Ca2+ sensitivity onto large-conductance (285 pS) K+ channels observed in inside-out patches. beta-Neuregulins have no effect on voltage-activated Ca2+ currents of CG neurons. These data support the hypothesis that beta-neuregulins mediate the trophic effects of preganglionic nerve terminals on the electrophysiological differentiation of developing CG neurons.

Posttranslational regulation of Ca(2+)-activated K+ currents by a target-derived factor in developing parasympathetic neurons.

Macroscopic IK[Ca is not expressed in normal levels in chick ciliary ganglion (CG) neurons prior to synapse formation with target tissues, or in neurons developing in vitro or in situ in the absence of target tissues. Here, two chick CG slo partial cDNAs encoding IK[Ca channels were isolated, cloned, and sequenced. Both slo transcripts were readily detected in developing CG neurons prior to or in the absence of target tissue interactions. When CG neurons developed in vitro in the presence of target tissue (iris) extracts, a normal whole-cell IK[Ca was expressed. These effects did not require protein synthesis, and the activity was detectable throughout the stages of synapse formation in the iris. The active component has an apparent molecular weight of 40-60 kDa.

The effects of innervation on the developmental expression of Ca(2+)-activated K+ currents in embryonic parasympathetic neurons are not activity-dependent.

Chronic blockade of synaptic transmission in ovo using mecamylamine, a neuronal nicotinic receptor antagonist, caused a large increase in naturally occurring cell death in the embryonic chick ciliary ganglion. However, the Ca(2+)-activated K+ currents in embryonic day 13 mecamylamine-treated ciliary ganglion neurons were indistinguishable from those of saline-treated controls. Therefore, the trophic effect of preganglionic innervation on the developmental expression of Ca(2+)-activated K+ current is not dependent upon intact nicotinic cholinergic synaptic transmission and may instead be mediated by a nerve terminal-derived differentiation factor.