Oestradiol Stimulates the Release of AVP and GnRH from the Ewe Hypothalamus In Vitro 1

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The present study aims at ascertaining the influence of alpha(1)-adrenoreceptors on arginine vasopressin (AVP) release in vitro and determine whether E(2) modulates the alpha(1)-adrenoreceptor and AVP interaction. Ten minutes after ewe killing, sagittal midline hypothalamic slices (from the anterior preoptic area to the mediobasal hypothalamus with the median eminence, 2 mm thick, 2 per sheep) were dissected, placed in oxygenated minimum essential media-alpha (MEM-alpha) at 4 degrees C and within 2 h were singly perifused at 37 degrees C with oxygenated MEM-alpha (pH 7.4; flow rate 0.15 ml/min), either with or without E(2) (24 pg/ml). After 4 h equilibration, 10 min fractions were collected for 4 h interposed with 10 min exposure at 60 min to a specific alpha(1)-adrenoreceptor agonist or antagonist at various doses (0.1-10 mm). At the end of all perifusions, slices responded to KCl (100 mm) with AVP efflux (p < 0.05). Release of AVP was enhanced (p < 0.05) by the alpha(1)-adrenoreceptor agonist (methoxamine 10 mm; no E(2), n = 7 perifusion chambers: from 14.3 +/- 2.7 to 20.9 +/- 3.9, with E(2), n = 10: from 10.7 +/- 1.2 to 18.4 +/- 3.4 pg/ml) or the antagonist (thymoxamine 10 mm; no E(2), n = 5: from 9.5 +/- 3.1 to 30.4 +/- 6.0, with E(2), n = 10: from 10.8 +/- 0.9 to 39.1 +/- 6.3 pg/ml). With the agonist, the response occurred only at 80 min (p < 0.05) both in the presence and absence of E(2). Whereas, after the antagonist, values were higher (p < 0.05) throughout the post-treatment period (80-170 min) without E(2), but declined by 150 min in the presence of E(2). Furthermore, the response to the alpha(1)-adrenoreceptor antagonist was greater (p < 0.05; 90-140 min) than the agonist only in the presence of E(2). In conclusion, these results reveal direct alpha(1)-adrenoreceptor-mediated control of the hypothalamic AVP neuronal system which is modulated by E(2).

“…Basal release of AVP from the hypothalamic slices was augmented by E 2 confirming our previous in vitro data (Ghuman et al. 2006) in agreement with in vivo studies in sheep (Wood et al.…”

Section: Discussionsupporting

confidence: 91%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 90%

Section: Introductionmentioning

confidence: 99%

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Noradrenergic Control of Arginine Vasopressin Release from the Ewe Hypothalamus In Vitro: Sensitivity to Oestradiol

Ghuman¹,

Prabhakar²,

Smith³

et al. 2008

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γ‐Amino Butyric Acid Control of Arginine Vasopressin Release from the Ewe Hypothalamus In Vitro: Sensitivity to Oestradiol

Ghuman

Prabhakar

Smith

et al. 2007

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The present study aims to ascertain the influence of gamma-amino butyric acid (GABA)(A or B) receptors on arginine vasopressin (AVP) release in vitro and determine whether E(2) modulates GABA-AVP interaction. Within 10 min of ewe killing, saggital midline hypothalamic slices (from the anterior preoptic area to the mediobasal hypothalamus along with the median eminence, 2-mm thick, two per ewe) were dissected, placed in oxygenated minimum essential media (MEM)-alpha at 4 degrees C and within 2 h were singly perifused at 37 degrees C with oxygenated MEM-alpha (pH 7.4; flow rate 0.15 ml/min), either with or without E(2) (24 pg/ml). After 4-h equilibration, 10-min fractions were collected for 4 h interposed with a 10-min exposure at 60 min to a specific GABA(A or B) receptor agonist or antagonist at various doses (0.1-10 mm). GABA(A) (muscimol; no E(2), n = 7 perifusion chambers, with E(2), n = 11) or GABA(B) (baclofen; no E(2), n = 8, with E(2), n = 15) agonists (10 mm) did not influence AVP concentrations. However, AVP release increased (p < 0.05) 20-30 min after exposure to 10 mm GABA(A or B) antagonists (bicuculline, no E(2), n = 7: from 4.6 +/- 0.7 to 33.0 +/- 0.4, with E(2), n = 17: from 11.9 +/- 1.4 to 32.8 +/- 6.0; CGP52432, with E(2), n = 14: from 14.0 +/- 2.6 to 28.8 +/- 3.9 pg/ml). At the end of the collection period, hypothalamic slices responded to KCl (100 mm) with AVP efflux (p < 0.05). GABA(B) but not GABA(A) antagonist-stimulated AVP release was enhanced in the presence of E(2). In summary, AVP release is under the inhibitory influence of GABA input with further potentiation by E(2) through GABA(B) receptors in vitro.

GABA Control of GnRH Release from the Ewe Hypothalamus In Vitro: Sensitivity to Oestradiol

Ghuman

Jones

Prabhakar

et al. 2008

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The present study examines the involvement of GABA(A or B) receptors in gonadotrophin-releasing hormone (GnRH) release in vitro and determines whether oestradiol modulates gamma-aminobutyric acid (GABA)-GnRH interaction. Within 10 min after ewe killing, hypothalamic slices were dissected and placed in oxygenated Minimum Essential Media (MEM)-alpha at 4 degrees C; within 2 h, slices were singly perifused at 37 degrees C with oxygenated MEM-alpha (0.15 ml/min), with or without oestradiol (24 pg/ml). After 4 h equilibration, fractions were collected for 4 h interposed with a 10 min exposure to specific GABA(A or B) receptor ligands (0.1-10 mM). The GABA(A or B) agonists (muscimol or baclofen) did not greatly influence GnRH release. However, GnRH increased (p < 0.05) after exposure to 10 mM GABA(A or B) antagonists (bicuculline or CGP52432, respectively). The GABA(A) antagonist stimulated greater sustained GnRH release (p < 0.05) in the absence of oestradiol than in its presence. The bioactivity of the released GnRH was studied using a hypothalamus-pituitary sequential double-chamber perifusion. Only after exposure of hypothalamic slices to the GABA(A) antagonist, did the hypothalamic eluate stimulate luteinizing hormone release from pituitary fragments (p < 0.05) confirming that the GABA(A) antagonist stimulated release of biologically active GnRH. In summary, GnRH release from the hypothalamus is predominantly under GABA(A) receptor inhibitory control and this is attenuated in the presence of oestradiol.