Insights into GABA receptor signalling in TM3 Leydig cells

gamma-Aminobutyric acid (GABA) is an emerging signalling molecule
in endocrine organs, since it is produced by endocrine cells and
acts via GABA(A) receptors in a paracrine/autocrine fashion.
Testicular Leydig cells are producers and targets for GABA. These
cells express GABA(A) receptor subunits and in the murine Leydig
cell line TM3 pharmacological activation leads to increased
proliferation. The signalling pathway of GABA in these cells is not
known in this study. We therefore attempted to elucidate details of
GABA(A) signalling in TM3 and adult mouse Leydig cells using
several experimental approaches. TM3 cells not only express GABA(A)
receptor subunits, but also bind the GABA agonist {[}H-3] muscimol
with a binding affinity in the range reported for other endocrine
cells (K-d = 2.740 +/- 0.721 nM). However, they exhibit a low B-max
value of 28.08 fmol/mg protein. Typical GABA(A) receptor-associated
events, including Cl- currents, changes in resting membrane
potential, intracellular Ca2+ or cAMP, were not measurable with the
methods employed in TM3 cells, or, as studied in part, in primary
mouse Leydig cells. GABA or GABA(A) agonist isoguvacine treatment
resulted in increased or decreased levels of several mRNAs,
including transcription factors (c-fos, hsf-1, egr-1) and cell
cycle-associated genes (Cdk2, cyclin D1). In an attempt to verify
the cDNA array results and because egr-1 was recently implied in
Leydig cell development, we further studied this factor. RT-PCR and
Western blotting confirmed a time-dependent regulation of egr-1 in
TM3. In the postnatal testis egr-1 was seen in cytoplasmic and
nuclear locations of developing Leydig cells, which bear GABA(A)
receptors and correspond well to TM3 cells. Thus, GABA acts via an
untypical novel signalling pathway in TM3 cells. Further details of
this pathway remain to be elucidated. Copyright (c) 2005 S. Karger
AG, Basel