ABSTRACT
A synRas mouse model was used expressing constitutively activated Ha-Ras (Val12 mutation) in neurons to investigate the role of Ras-MAPkinase signalling for neuronal connectivity in adult brain. Expression of the transgene in the cortex of these mice starts after neuronal differentiation is completed and allows to directly investigate the effects of enhanced Ras activity in differentiated neurons. Activation of Ha-Ras induced an increase in soma size which was sensitive to MEK inhibitor in postnatal organotypic cultures. Adult cortical pyramidal neurons showed complex structural rearrangements associated with an increased size and ramification of dendritic arborization. Dendritic spine density was elevated and correlated with a twofold increase in number of synapses. In acute brain slices of the somatosensory and of the visual cortex, extracellular field potentials were recorded from layer II/III neurons. The input-output relation of synaptically evoked field potentials revealed a significantly higher basal excitability of the transgenic mice cortex compared to wild-type animals. In whole cell patch clamp preparations, the frequency of AMPA receptor-mediated spontaneous excitatory postsynaptic currents was increased while the ratio between NMDA and AMPA-receptor mediated signal amplitude was unchanged. A pronounced depression of paired pulse facilitation indicated that Ras contributes to changes at the presynaptic site. Furthermore, synRas mice showed an increased synaptic long-term potentiation, which was sensitive to blockers of NMDA-receptors and of MEK. We conclude that neuronal Ras is a common switch of plasticity in adult mammalian brain sculpturing neuronal architecture and synaptic connectivity in concert with tuning synaptic efficacy.
Subject(s)
Cerebral Cortex/cytology , Mitogen-Activated Protein Kinase Kinases/physiology , Pyramidal Cells/physiology , Synapses/physiology , ras Proteins/metabolism , 2-Amino-5-phosphonovalerate/pharmacology , Animals , Animals, Newborn , Axons/ultrastructure , Butadienes/pharmacology , Cell Count/methods , Cell Size/genetics , Cell Size/physiology , Cells, Cultured , Dendrites/ultrastructure , Dose-Response Relationship, Drug , Electric Stimulation , Enzyme Inhibitors/pharmacology , Excitatory Amino Acid Agonists/pharmacology , Excitatory Amino Acid Antagonists/pharmacology , Excitatory Postsynaptic Potentials/drug effects , Excitatory Postsynaptic Potentials/physiology , Histocytochemistry/methods , Immunohistochemistry/methods , In Vitro Techniques , Long-Term Potentiation/drug effects , Long-Term Potentiation/physiology , Mice , Mice, Transgenic/genetics , Microscopy, Electron/methods , Nitriles/pharmacology , Patch-Clamp Techniques/methods , Pyramidal Cells/ultrastructure , Quinoxalines/pharmacology , Rats , Synapses/ultrastructure , Synaptic Transmission/drug effects , Synaptic Transmission/physiology , Synaptophysin/metabolism , Time Factors , alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid/pharmacology , ras Proteins/geneticsABSTRACT
The small G protein p21Ras is a critical molecular switch for relaying neurotrophic actions and is essential for normal functioning and plasticity of the nervous system. In this study, the morphogenetic effects of p21Ras were investigated on neurons in vivo. Morphological changes of layers II/III and Vb commissural pyramidal neurons of the primary somatosensory cortex were analyzed in transgenic mice expressing permanently active p21H-RasVal12 in postmitotic neurons. Pyramidal cells were retrogradely labelled with biotinylated dextran amine and subsequently traced using Neurolucida. Compared with wild-type mice, transgenic animals showed a significant increase in the surface area and volume of basal dendrites on the proximal and intermediate segments in layers II/III and on further distal segments in layer V. In addition, the surface area and volume of the trunk and of the proximal segments of oblique branches of apical dendrites were enlarged in both layers. Sholl analyses of basal and apical dendrites showed a significant increase in dendritic complexity of layer V neurons. A positive correlation was observed between the size of the basal dendrite and the neuronal soma size in the transgenic population, indicating that growth-promoting effects of p21H-RasVal12 affect both cellular compartments in parallel. However, the dendritic surface correlated with the number of tips and dendritic stem diameter in both wild-type and transgenic populations, demonstrating that these relations represent rather conservative design principles in dendritic morphology. The data presented here suggest an important role of p21Ras-dependent signaling in the final differentiation and maintenance of dendritic morphology.
