IMBECU   20882
INSTITUTO DE MEDICINA Y BIOLOGIA EXPERIMENTAL DE CUYO
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
PUBERTY AND NEUROSTEROIDS: EFFECT OF ALLOPREGNANOLONE ON GABA AND GLUTAMATE RELEASE AND DIFFERENTIAL HYPOTHALAMIC EXPRESSION AND ACTIVITY OF 3α-HOR IN FEMALE RATS
Autor/es:
GIULIANI F.; ESCUDERO C.; CASAS S.; GARCÍA S.; BAZZOCCHINI V.; YUNES R.; CABRERA R.
Lugar:
Viña del Mar
Reunión:
Workshop; US-LATINOAMERICAN WORKSHOP IN NEUROENDOCRINOLOGY; 2011
Resumen:
Introduction: The onset of puberty in female rats is normally triggered by an early
pulsatile GnRH release and is evident when vaginal opening occurs. GnRH release
appears to be regulated by transynaptic and glial inputs to the GnRH neuronal network,
like glutamatergic and GABAergic inputs. Such neuronal activity may be regulated, in
turns, by neurosteroids.
Allopregnanolone (Allo) is one of the best characterized neurosteroids. It is synthesized
by the enzyme 3α-hydroxysteroid oxydoreductase (3αHOR) from dihydroprogesterone.
Previous works of our laboratory revealed that Allo stimulates GnRH release through
NMDA receptors modulation in medial basal hypothalamus (MBH) and anterior
preoptic area (APOA) of adult rats. However, little is known about allopregnanolone
effects in an earlier stage such as puberty.
Objetives: 1) To determine the gene expression of 3αHOR in MBH/APOAs of
prepubertal rats (PP), rats on the day of vaginal opening (VO) and pubertal rats (P), 2)
to measure the 3αHOR enzymatic activity in MBH/APOAs of PP, VO and P rats and 3)
to determine the effect of Allo on GABA and glutamate release from MBH/APOA
slices of PP, VO and P rats.
Methods: 1) RNA was isolated from MBH/APOAs from PP (30d) (n=6), VO (39d)
(n=5) and P (55d) (n=6) rats. Multiplex RT-PCR was made by using primers for
3αHOR and ciclophiline (housekeeping gene). Relative optic density was obtained after
electrophoresis and etidium bromide dying. 2) MBH/APOAs of rats of the three
experimental groups (n=8 for each group) were homogenized and ultracentrifuged.
Then, citoplasmatic fraction was isolated to assay 3αHOR enzymatic activity by
spectrophotometric measure of oxidation of NADPH after addition of
dihydroprogesterone. 3) MBH/APOA slices of rats in the three experimental groups
were assayed for K+ 28mM evoked [3H]-GABA and [3H]-Glutamate release. They were
superfused with Krebs Ringer Bicarbonate Glucose (KRBG) buffer as vehicle or
alloprengnanolone 120 nM. Results were compared by ANOVA1 and Turkeys post
test, and differences with p