ABSTRACT
Ras-GRF1 is a neuron-specific guanine nucleotide exchange factor for Ras proteins. Mice lacking Ras-GRF1 (-/-) are severely impaired in amygdala-dependent long-term synaptic plasticity and show higher basal synaptic activity at both amygdala and hippocampal synapses (Brambilla et al., 1997). In the present study we investigated the effects of Ras-GRF1 deletion on hippocampal neuronal excitability. Electrophysiological analysis of both primary cultured neurons and adult hippocampal slices indicated that Ras-GRF1-/- mice displayed neuronal hyperexcitability. Ras-GRF1-/- hippocampal neurons showed increased spontaneous activity and depolarized resting membrane potential, together with a higher firing rate in response to injected current. Changes in the intrinsic excitability of Ras-GRF1-/- neurons can entail these phenomena, suggesting that Ras-GRF1 deficiency might alter the balance between ionic conductances. In addition, we showed that mice lacking Ras-GRF1 displayed a higher seizure susceptibility following acute administration of convulsant drugs. Taken together, these results demonstrated a role for Ras-GRF1 in neuronal excitability.
Subject(s)
Action Potentials/physiology , Hippocampus/metabolism , Pyramidal Cells/metabolism , ras-GRF1/deficiency , Action Potentials/drug effects , Animals , Electric Stimulation , Excitatory Amino Acid Antagonists/pharmacology , Excitatory Postsynaptic Potentials/drug effects , Excitatory Postsynaptic Potentials/physiology , Female , Genetic Predisposition to Disease/genetics , Glutamate Decarboxylase/metabolism , Hippocampus/cytology , Hippocampus/drug effects , Isoenzymes/metabolism , Male , Mice , Mice, Knockout , Nerve Net/cytology , Nerve Net/drug effects , Nerve Net/metabolism , Patch-Clamp Techniques , Pyramidal Cells/cytology , Pyramidal Cells/drug effects , Seizures/chemically induced , Seizures/genetics , Seizures/metabolism , Synaptic Transmission/drug effects , Synaptic Transmission/physiology , Synaptophysin/metabolism , Tetrodotoxin/pharmacology , ras-GRF1/geneticsABSTRACT
Monoaminergic G protein-coupled receptors (GPCRs) are highly expressed in the CNS at the cerebrocortical level, where they support a variety of behavioural responses. To elucidate possible intracellular signalling pathways coupled to these receptors, we have studied their ability to activate extracellular signal-regulated kinases (ERKs) in cultured cortical neurons. An increase in ERK activity was observed after stimulation of neurons with dopamine or serotonin, and with agonists selective for various GPCRs. In addition, ERK activation was also observed following treatment with phorbol dibutyrate (PdBu) and forskolin, activators of protein kinase C (PKC) and protein kinase A (PKA), respectively. Concomitant with ERK activation, all the monoaminergic agonists tested also increased the level of active Ras (Ras-GTP). Surprisingly, Ras activation was also observed after activation of cAMP pathway, and this effect was at least in part mediated by PKA. Ras activation by cAMP was unique for neurons, since in PC12 cells forskolin caused activation of ERK but did not increase Ras-GTP level. These results highlight the relevance of Ras as a target for multiple signalling cascades leading to activation of the ERK pathway in neurons.
