Quantitative reverse transcription and polymerase chain reaction analysis of oxytocin and vasopressin receptor mRNAs in the rat uterus near parturition.

Oxytocin receptors (OT-R) are known to be involved in the course of labor since a massive increase in OT-binding sites is observed in the uterus just before parturition. Vasopressin (AVP)-binding sites have also been observed and have been shown to mediate uterotonic responses. To determine exactly which subtypes of OT/AVP receptors are expressed in the rat uterus near parturition, we carried out absolute quantitations of the neurohypophysial hormone receptor (OT-R and the vasopressin receptors V1a-R, V1b-R and V2-R) mRNAs with an assay based on reverse transcription-polymerase chain reaction (RT-PCR) using in vitro transcribed mutated cRNAs as internal standards. The number of mRNA molecules/ng of total RNA was 35 +/- 6, 220 +/- 33 and 39 +/- 9 for OT-R (P < 0.01) and 16 +/- 1, 25 +/- 8 and 31 +/- 5 for V1a-R (P > 0.05) on day (D) 21, 22 and 23 of gestation (post-parturient), respectively. We did not detect V1b-R and V2-R mRNAs in the pregnant uterus. Therefore, the heterogeneity of OT and AVP receptors in the rat uterus can only be assigned to the presence of OT-R and V1a-R neurohypophysial hormone receptor subtypes, whereas V1b-R and V2-R can not be invoked. Only OT-R mRNA levels change in the uterus near parturition.

Comparison of vasopressin and oxytocin receptors in the rat uterus and vascular tissue.

Several studies indicate that oxytocin and vasopressin receptors in the human uterus are heterogeneous. We have investigated whether oxytocin and vasopressin bind to separate receptors in the day 21 and day 22 pregnant rat uterus and whether uterine vasopressin receptors are the same as the vascular V1A subtype. In isolated organ bath experiments we showed that the potency of d(CH2)5[Tyr(Me)2]vasopressin to inhibit vasopressin contraction in rat aorta was different from that in the day 21 pregnant uterus. Saturation curves of [3H]vasopressin in membranes from cultured aortic myocytes and pregnant uterus were linear and yielded the same 1 nM Kd values. However, the potency of d(CH2)5[Tyr(Me)2]vasopressin and of [Thr4,Gly7]oxytocin at antagonizing [3H]vasopressin confirmed the differences between the vascular smooth muscle and uterine vasopressin receptor. The peptides had respectively higher and lower affinity for aortic cell sites than for uterine sites. It was more difficult to distinguish pharmacological differences for oxytocin and vasopressin receptors in the uterus. On day 22, the high affinity of [Thr4,Gly7]oxytocin and oxytocin for both [3H]oxytocin and [3H]vasopressin binding sites was consistent with the notion that the uterus expresses essentially oxytocin receptors at this stage of gestation. However, oxytocin, vasopressin and three analogs showed a different potency for inhibiting [3H]oxytocin and [3H]vasopressin binding on day 21 versus day 22 of gestation. We conclude that in the rat uterus vasopressin binds to a receptor that is different from the vascular V1A subtype. Also, the binding sites for [3H]vasopressin and [3H]oxytocin on day 21 uterus membranes do not resemble the classical oxytocin receptor as described in the literature suggesting that on day 21 vasopressin and oxytocin bind in the uterus to a receptor that might be different from those currently characterized.

Effects of nomegestrol acetate on spontaneous and sulprostone-induced uterine contractions in pregnant cynomolgus monkeys monitored by telemetry.

OBJECTIVE: Our purpose was to study the effects of the progestomimetic compound nomegestrol acetate on spontaneous and sulprostone-induced uterine contractility in pregnant cynomolgus monkeys. STUDY DESIGN: Intrauterine pressure was continuously monitored with use of an implanted intraamniotic catheter and a telemetric pressure transmitter from day 115 to 135 of gestation (term = 165 days). After surgery the animals received either nomegestrol acetate (5 mg per day orally, n = 3) or vehicle only (controls, n = 3). The intramuscular prostaglandin E2 analog sulprostone (25 micrograms) was administered as a single injection 10 days after amniotic catheter implantation. Spontaneous and sulprostone-induced uterine contractions were compared between nomegestrol acetate- and vehicle-treated animals. RESULTS: The frequency of spontaneous uterine contractions in control animals demonstrated a 24-hour pattern with a minimum at 12 hours and a maximum at 0 hours. The frequency of spontaneous contractions did not differ between nomegestrol acetate- and vehicle-treated animals. Sulprostone induced an increase in both the frequency and amplitude of contractions, reaching a maximum 12 hours after injection and fading out after 24 hours in vehicle-treated animals. In animals receiving nomegestrol acetate, the frequency of contractions increased moderately and transiently for a total duration of 6 hours only and returned to control levels thereafter. CONCLUSION: Nomegestrol acetate significantly decreases the contractile response of the pregnant uterus induced by the prostaglandin E2 analog sulprostone.

beta2-adrenergic but not vasopressin V2 receptor mRNAs are expressed in the stria vascularis of the rat inner ear.

Activation of beta2-adrenergic receptors (beta2-AR) and of V2-vasopressin receptors (V2-VR) has been shown to stimulate cAMP production in the stria vascularis. Expression of these receptors in this epithelial structure has never been investigated at the mRNA level. A quantitative assay based on the reverse transcriptase-polymerase chain reaction (RT-PCR) method was developed to study the expression of beta2-AR, vasopressin (V1a,V1b, V2) and oxytocin receptors in rat microdissected stria vascularis fragments. Steady-state mRNA levels were measured using mutant cRNAs as internal standards. We consistently found evidence of the expression of beta2-AR transcripts in the stria vascularis; however, no such evidence of V2-VR mRNA expression was found in studies of the same samples. As our method is sensitive enough to detect 200 mRNA molecules, V2-VR mRNA is thought to be present, if at all, at a concentration that is 40 times lower than that of the beta2-AR transcripts. V1a-VR mRNA was found to be present in trace amounts and is the only subtype of the vasopressin-oxytocin receptor family expressed in this epithelium. These data demonstrate, at the mRNA level, the expression of beta2-AR in the stria vascularis, and indicate that V2-VR transcripts are not present in this structure.