GABA(A) receptors present higher affinity and modified subunit composition in spinal motor neurons from a genetic model of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis is a neurodegenerative disease characterized by the selective degeneration of motor neurons in the spinal cord, brainstem and cerebral cortex. In this study we have analysed the electrophysiological properties of GABA(A) receptors and GABAA alpha1 and alpha2 subunits expression in spinal motor neurons in culture obtained from a genetic model of ALS (G93A) and compared with transgenic wild type SOD1 (SOD1) and their corresponding non transgenic litter mates ( Control). Although excitotoxic motor neuron death has been extensively studied in relation to Ca2+-dependent processes, strong evidence indicates that excitotoxic cell death is also remarkably dependent on Cl- ions and on GABA(A) receptor activation. In this study we have analysed the electrophysiological properties of GABAA receptors and the expression of GABA(A) alpha 1 and alpha 2 subunits in cultured motor neurons obtained from a genetic model of amyotrophic lateral sclerosis (G93A) and compared them with transgenic wild-type Cu, Zn superoxide dismutase and their corresponding non-transgenic littermates (Control). In all tested motor neurons, the application of gamma-aminobutyric acid (GABA) (0.5-100 mu M) evoked an inward current that was reversibly blocked by bicuculline (100 mu M), thus indicating that it was mediated by the activation of GABAA receptors. Our results indicate that the current density at high GABA concentrations is similar in control, Cu, Zn superoxide dismutase and G93A motor neurons. However, the dose-response curve significantly shifted toward lower concentration values in G93A motor neurons and the extent of desensitization also increased in these neurons. Finally, multiplex single-cell real-time polymerase chain reaction and immunofluorescence revealed that the amount of GABA(A) alpha 1 subunit was significantly increased in G93A motor neurons, whereas the levels of alpha 2 subunit were unchanged. These data show that the functionality and expression of GABA(A) receptors are altered in G93A motor neurons inducing a higher Cl- influx into the cell with a possible consequent neuronal excitotoxicity acceleration.