Regulation of N- and L-Type Ca²⁺Channels in Adult Frog Sympathetic Ganglion B Cells by Nerve Growth Factor In Vitro and In Vivo

Abstract

To examine mechanisms responsible for the long-term regulation of Ca2+-channels in an adult neuron, changes in whole cell Ba2+ current (IBa) were examined in adult bullfrog sympathetic ganglion B cells in vitro. Cells were cultured at low density in defined, serum free medium. After 15 days, total IBa was similar to the initial value, whereas IBa density was reduced by approximately 36%, presumably due to an increase in neuronal surface area. By contrast, IBa density remained constant after 6-15 days in the presence of murine beta-NGF (200 ng/ml), and total IBa was almost doubled. Inclusion of cytosine arabinoside (Ara-C; 10 microM) to inhibit proliferation of nonneuronal cells, did not affect the survival of neurons in the absence of nerve growth factor (NGF) nor did it attenuate IBa. Ara-C did not prevent the effect of NGF on IBa. There were three independent components to the action of NGF; during 6-9 days, it increased omega-conotoxin-GVIA-sensitive N-type IBa (IBa,N); increased nifedipine-sensitive L-type IBa (IBa,L) and decreased inactivation of the total Ba2+ conductance (gBa). The latter effect involved a selective decrease in the amplitude of one of the four kinetic components that describe the inactivation process. Total IBa was also 55.8% larger than control in the somata of B cells acutely dissociated from leopard frogs that had received prior subcutaneous injections of NGF. By contrast, injection of NGF antiserum decreased total IBa by 29.4%. There was less inactivation of gBa in B cells from NGF-injected animals than in cells from animals injected with NGF antiserum (P < 0.001). These data suggest that NGF-like molecule(s) play(s) a role in the maintenance of IBa in an adult amphibian sympathetic neuron; the presence of NGF may allow the neuron to maintain a constant relationship between cell size and current density. They also show that IBa inactivation in an adult neuron can be modulated in a physiologically relevant way by an extracellular ligand.

Exogenous nerve growth factor attenuates opioid-induced inhibition of voltage-activated Ba2+ currents in rat sensory neurons

“…For example, NGF has been shown to rapidly increase capsaicin-induced currents in primary afferent nociceptors (Shu and Mendell, 2001) as well as increase currents through VGCC in some cells (Wildering et al, 1995;Jia et al, 1999) within a few minutes. On the other hand, TRPV1 receptors can also be upregulated by NGF with a time course of hours to days (Nicholas et al, 1999;Ji et al, 2002), as can VGCC (Lesser and Lo, 1995;Levine et al, 1995;Fitzgerald and Dolphin, 1997;Lei et al, 1997). The lack of a rapid effect on opioidmediated inhibition of I Ba when NGF was applied acutely in the present study suggests that this effect was not directly mediated by protein phosphorylation or dephosphorylation, but involved a longer term, possibly transcriptional event.…”

“…NGF has been shown to upregulate the TRPV1 channel, one of the molecular transducers of noxious heat which is also activated by capsaicin, and to increase capsaicin-induced currents in nociceptors (Nicholas et al, 1999;Shu and Mendell, 2001;Ji et al, 2002), thereby contributing to thermal hyperalgesia. NGF also increases currents through voltage-gated calcium channels (VGCC) in a wide variety of cells (Lesser and Lo, 1995;Levine et al, 1995;Wildering et al, 1995;Lei et al, 1997;Jia et al, 1999) including sensory neurons (Fitzgerald and Dolphin, 1997). Upregulation of VGCC may enhance nociceptor excitability, and promote the release of excitatory substances from nociceptor terminals.…”

Differential Neurotrophic Regulation of Sodium and Calcium Channels in an Adult Sympathetic Neuron

“…Although "molecular" approaches such as RNA interference (Holen and Mobbs 2004) or introduction of dominant negative or constitutively active forms of ras (Fitzgerald 2000) might be considered, the lack of information on the Rana catesbeiana genome and the fact that suitable reagents are not readily available makes this approach difficult or infeasible. Since this study is a continuation of an extensive series of physiological studies (Ford et al 2003a;Jassar et al 1993;Lei et al 1997Lei et al , 1998Lei et al , 2001Petrov et al 2001) on Rana catesbeina, it is not appropriate to change to a mammalian (Fitzgerald 2000;Fitzgerald and Dolphin 1997) or cell line system even though they are clearly more amenable to molecular biological approaches (Black et al 2003).…”

“…Although LHRH exclusively increases I Ca,N with no effect on activation or inactivation kinetics (Ford et al 2003a), the effects of NGF are more complex and involve an increase in I Ca,L , decreased inactivation of total I Ca , and an increase in I Ca,N (Lei et al 1997). Mechanisms additional to ras-MAPK may be involved in NGF effects on I Ca,L and inactivation processes.…”

“…Various neurotrophic factors have therefore been studied as important determinants of ion channel expression in adult neurons and in cell lines (Chalazonitis et al 1987;Fanger et al 1997;Khorkova and Golowasch 2007;Levine et al 1995;Pollock and Rane 1996;Pollock et al 1990). Target-derived nerve growth factor (NGF) is responsible for the maintenance of tetrodotoxin (TTX)-sensitive and TTX-insensitive sodium currents (I Na ) and of N-and L-type calcium currents (I Ca,N and I Ca,L ) in B-neurons of adult bullfrog sympathetic ganglia (BFSG) (Lei et al 1997(Lei et al , 1998(Lei et al , 2001Petrov et al 2001). In serum-free, defined medium culture, NGF-induced increases in I Ca are initiated via the mitogen-activated protein kinase (MAPK) pathway (Lei et al 1998), whereas the transduction process underlying the NGF-induced increase in I Na remains to be determined.…”