Recombinant GABA(C) receptors expressed in rat hippocampal neurons after infection with an adenovirus containing the human rho1 subunit.
1. A recombinant adenovirus was generated with the human rho1 GABA(C) receptor subunit (adeno-rho). Patch-clamp and antibody staining were employed to confirm functional expression of recombinant rho1 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 rho1 GABA(C) receptor antibodies revealed that recombinant rho1 receptors were expressed in HEK293 and 911 cells after adeno-rho infection and exhibited properties similar to those of rho1 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-rho 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 rho1 receptors could be expressed in cultured hippocampal slices after adeno-rho 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.
Filippova
N
N
Department of Neurobiology, University of Alabama at Birmingham School of Medicine, 1719 Sixth Avenue South CIRC 410, Birmingham, AL 35294-0021, USA.
Sedelnikova
A
A
Tyler
W J
WJ
Whitworth
T L
TL
Fortinberry
H
H
Weiss
D S
DS
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D002460:Cell Line; D004553:Electric Conductivity; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D009474:Neurons / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D018079:Receptors, GABA / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D018080:Receptors, GABA-B / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D020033:Protein Isoforms; D018079:Receptors, GABA; D018080:Receptors, GABA-B; D011994:Recombinant Proteins