New insights on the role of gephyrin in regulating both phasic and tonic GABAergic inhibition in rat hippocampal neurons in culture.

Gephyrin is a tubulin-binding protein that acts as a scaffold for clustering glycine and GABA(A) receptors at postsynaptic sites. In this study, the role of gephyrin on GABA(A) receptor function was assessed at the post-translational level, using gephyrin-specific single chain antibody fragments (scFv-gephyrin). When expressed in cultured rat hippocampal neurons as a fusion protein containing a nuclear localization signal, scFv-gephyrin were able to remove endogenous gephyrin from GABA(A) receptor clusters. Immunocytochemical experiments revealed a significant reduction in the number of synaptic gamma2-subunit containing GABA(A) receptors and a significant decrease in the density of the GABAergic presynaptic marker vesicular GABA transporter (VGAT). These effects were associated with a slow down of the onset kinetics, a reduction in the amplitude and in the frequency of miniature inhibitory postsynaptic currents (mIPSCs). The quantitative analysis of current responses to ultrafast application of GABA suggested that changes in onset kinetics resulted from modifications in the microscopic gating of GABA(A) receptors and in particular from a reduced entry into the desensitized state. In addition, hampering gephyrin function with scFv-gephyrin induced a significant reduction in GABA(A) receptor-mediated tonic conductance. This effect was probably dependent on the decrease in GABAergic innervation and in GABA release from presynaptic nerve terminals. These results indicate that gephyrin is essential not only for maintaining synaptic GABA(A) receptor clusters in the right position but also for regulating both phasic and tonic inhibition.

Comparison between low-level 50 Hz and 900 MHz electromagnetic stimulation on single channel ionic currents and on firing frequency in dorsal root ganglion isolated neurons.

Alteration of membrane surface charges represents one of the most interesting effects of the electromagnetic exposure on biological structures. Some evidence exists in the case of extremely low frequency whereas the same effect in the radiofrequency range has not been detected. Changes in transmembrane voltages are probably responsible for the mobilization of intracellular calcium described in some previous studies but not confirmed in others. These controversial results may be due to the cell type under examination and/or to the permeability properties of the membranes. According to such a hypothesis, calcium oscillations would be a secondary effect due to the induced change in the membrane voltage and thus dependent on the characteristics of ionic channels present in a particular preparation. Calcium increases could suggest more than one mechanism to explain the biological effects of exposure due to the fact that all the cellular pathways using calcium ions as a second messenger could be, in theory, disturbed by the electromagnetic field exposure. In the present work, we investigate the early phase of the signal transmission in the peripheral nervous system. We present evidence that the firing rate of rat sensory neurons can be modified by 50/60 Hz magnetic field but not by low level 900 MHz fields. The action of the 50/60 Hz magnetic field is biphasic. At first, the number of action potentials increases in time. Following this early phase, the firing rate decreases more rapidly than in control conditions. The explanation can be found at the single-channel level. Dynamic action current recordings in dorsal root ganglion neurons acutely exposed to the electromagnetic field show increased functionality of calcium channels. In parallel, a calcium-activated potassium channel is able to increase its mean open time.