Influence of Surgical Trauma in the Nasal Cavity on the Expression of p53 Protein in the Hippocampus of Rats.

The aim of the study was to determine the role of the stress effect of septoplasty modeling on p53 protein expression in the hippocampus of rats under conditions of sensory olfactory deprivation. Simulation of septoplasty was carried out on 30 sexually mature male rats. A quantitative assessment of the apoptosis of neurons in the pyramidal layer of the hippocampus in the subfields CA1, CA2, CA3, and dentate gyrus (DG) on days 2, 4, and 6 after surgery was carried out. Histological sections were stained by the immunohistochemical method with antibodies to the p53 protein. An increase in the number of p53-positive neurons was noted in all subfields; the maximum increase in the number of apoptotic neurons was noted on day 4 after surgery. The stress effect of modeling septoplasty in rats, accompanied by sensory deprivation of the peripheral part of the olfactory analyzer, provoked the expression of p53 and the initiation of apoptosis mechanisms in various subfields of the hippocampus.

alpha1beta2delta, a silent GABAA receptor: recruitment by tracazolate and neurosteroids.

BACKGROUND AND PURPOSE: This study investigated the alpha(1)beta(2)delta isoform of the GABA(A) receptor that is presumably expressed in the forebrain. The functional and pharmacological properties of this receptor combination are largely unknown. EXPERIMENTAL APPROACH: We expressed alpha(1)beta(2)delta GABA(A) receptors in Xenopus laevis oocytes. GABA-activated currents, in the presence and absence of modulators, were recorded using the two-electrode voltage clamp technique. KEY RESULTS: The alpha(1)beta(2)delta isoform of the GABA(A) receptor exhibited an extremely small GABA-mediated current. Tracazolate increased the current amplitude evoked by a half-maximal concentration (EC(50)) of GABA by 59-fold. The maximum current was increased 23-fold in the presence of a saturating GABA concentration. Concomitant with the increase in the maximum, was a 4-fold decrease in the EC(50). Finally, a mutation in the second transmembrane domain of the delta subunit that increases receptor efficacy (L286S), eliminated the increase in the maximum GABA-activated current. The endogenous neurosteroid, tetrahydrodeoxycorticosterone (THDOC), also decreased the EC(50) and increased the maximum current amplitude, although to a lesser degree than that of tracazolate. CONCLUSIONS AND IMPLICATIONS: Taken all together, these findings indicate that the small GABA-mediated currents in the absence of the modulator are due to a low efficacy for activation. In the absence of modulators, alpha(1)beta(2)delta GABA receptors would be effectively silent and therefore contribute little to inhibition in the CNS. In the presence of tracazolate or endogenous neurosteroids however, this particular receptor isoform could exert a profound inhibitory influence on neuronal activity.

Regulation of recombinant gamma-aminobutyric acid (GABA)(A) and GABA(C) receptors by protein kinase C.

Activation of protein kinase C (PKC) by phorbol 12-myristate 13-acetate induced a continuous decrease in the gamma-aminobutyric acid (GABA)-activated current amplitude from recombinant GABA receptors (formed by rho1 or alphabetagamma subunits) expressed in Xenopus oocytes. This decline was due to internalization of receptors from the plasma membrane as confirmed by a decrease in surface fluorescence with green fluorescence protein-tagged receptors as well as a concomitant decrease in surface [(3)H]GABA binding. PMA specifically caused internalization of GABA receptors, but not neuronal acetylcholine receptors (alpha(7) or alpha(4)beta(2)), indicating the internalization was not a general, nonspecific phenomenon. Mutation of rho1 PKC phosphorylation sites, identified by in vitro phosphorylation, did not prevent GABA receptor internalization, nor did coexpression of the rho1 M3-M4 intracellular loop along with rho1 GABA receptors. It is likely that PKC-mediated phosphorylation of other proteins, rather than rho1 itself, was required for the internalization. Both rho1 and alphabetagamma receptors did not degrade after phorbol 12-myristate 13-acetate-induced internalization, but returned to the membrane surface within 24 h. These data suggest internalized receptors can exist in an intracellular compartment that can be delivered back to the plasma membrane. Thus, by regulating GABA receptor surface expression, PKC may play a key role in the regulation of GABA-mediated inhibition.

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.