Novel Actions of Neurosteroids on GABAA Receptors.
Comenencia Ortiz, Eydith.
Gamma-aminobutyric acid type A receptors (GABAARs) are the principal mediators of
inhibitory transmission in the mammalian central nervous system. GABAARs can be
localized at postsynaptic inhibitory specializations or at extrasynaptic sites. While
synaptic GABAARs are activated transiently following the release of GABA from
presynaptic vesicles, extrasynaptic GABAARs are activated ... read morecontinuously by resting GABA
concentrations and thus mediate tonic inhibition. In the hippocampus and thalamus,
extrasynaptic GABAARs are predominantly composed of alpha4 beta2/3 and delta subunits.
The tonic inhibitory currents mediated by extrasynaptic GABAARs control neuronal
excitability and the strength of synaptic transmission. However, the mechanisms by which
neurons control the functional properties of extrasynaptic GABAARs had not yet been
explored. Phosphorylation of residues within synaptic GABAAR subunits is critical for
the assembly, trafficking and cell surface stability of synaptic GABAAR subtypes. Here
we have identified serines 443 in the alpha4 and serines 408/409 in the beta3 subunit as
the principal sites for PKC phosphorylation in extrasynaptic GABAAR subtypes. Steroid
metabolites of progesterone and deoxycortisone (known as neurosteroids) have been shown
to be potent positive allosteric modulators of extrasynaptic GABAARs. However, the exact
mechanisms by which neurosteroids alter extrasynaptic GABAARs function are not well
understood. Previous experiments have suggested that Protein Kinase C (PKC) activity is
required for neurosteroid-mediated modulation of GABAARs. We show that the
deoxycortisone metabolite, Tetrahydrodeoxycorticosterone (THDOC) induces the
phosphorylation of the alpha4 GABAAR subunit on serine 443 (S443) dependent upon the
ability of this agent to activate PKC. We analyzed the functional significance of THDOC
induced phosphorylation of the alpha4 subunit using patch-clamp recording and TIRF
microscopy. Collectively these approaches revealed that THDOC increases the cell surface
stability of alpha4-containing GABAARs by promoting their insertion into the plasma
membrane, a phenomenon critically dependent on S443 in the alpha4 subunit and on
S408/409 within the beta3 subunit when co-expressed. Significantly, the ability of THDOC
to increase the expression levels of GABAARs was independent of the delta subunit and
glutamine 246 (Q246) in the alpha4 subunit, a site critical for allosteric neurosteroid
potentiation. Collectively we have identified a novel mechanism by which neurosteroids
can induce long-term changes in the expression levels of GABAARs via PKC dependent
phosphorylation of the alpha4 subunit.
Thesis (Ph.D.)--Tufts University, 2012.
Submitted to the Dept. of Neuroscience.
Advisor: Stephen Moss.
Committee: Jamie Maguire, Maribel Rios, Kathleen Dunlap, and Shelley Russek.
Keyword: Neurosciences.read less