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Recombinant GABA_C receptors expressed in rat hippocampal neurons after infection with an adenovirus containing the human 1 subunit

Natalia Filippova, Anna Sedelnikova, William J. Tyler, Terri L. Whitworth, Henry Fortinberry and David S. Weiss

1. A recombinant adenovirus was generated with the human 1 GABA_C receptor subunit (adeno-). Patch-clamp and antibody staining were employed to confirm functional expression of recombinant 1 receptors after infection of human embryonic kidney cells (HEK293 cell line), human embryonic retinal cells (911 cell line), dissociated rat hippocampal neurons and cultured rat hippocampal slices.
2. Standard whole-cell recording and Western blot analysis using 1 GABA_C receptor antibodies revealed that recombinant 1 receptors were expressed in HEK293 and 911 cells after adeno- infection and exhibited properties similar to those of 1 receptors after standard transfection.
3. Cultured rat hippocampal neurons (postnatal day (P)3-P5) did not show a native GABA_C-like current. After adeno- infection, however, a GABA_C-like current appeared in 70-90 % of the neurons.
4. Five days after infection, expression of GABA_C receptors in hippocampal neurons significantly decreased native GABA_A receptor currents from 1200 ± 300 to 150 ± 70 pA (n = 10). The native glutamate-activated current was unchanged.
5. Hippocampal slices (P8) did not show a native GABA_C-like current, although recombinant 1 receptors could be expressed in cultured hippocampal slices after adeno- infection.
6. These data indicate that an adenovirus can be used to express recombinant GABA_C receptors in hippocampal neurons. This finding could represent an important step towards the gene therapy of CNS receptor-related diseases.

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