Inga Neumann

Oxytocin and vasopressin release within the supraoptic and paraventricular nuclei of pregnant, parturient and lactating rats: A microdialysis study

The release of the nonapeptides oxytocin and vasopressin within the hypothalamic supraoptic and paraventricular nuclei was measured in 30-min microdialysates in conscious female rats in the last three days of pregnancy, during parturition, immediately after parturition and during suckling, all in the same rats, and in virgin controls. Nonapeptide release within the supraoptic and paraventricular nuclei was unchanged during late pregnancy compared to virgin rats, but intranuclear oxytocin and not vasopressin release was elevated during parturition (relative to late pregnancy, supraoptic nucleus: to 254%, paraventricular nucleus: to 300%; P < 0.01) and during suckling also on days 8-10 of lactation (relative to pre-suckling, supraoptic nucleus: to 407%, paraventricular nucleus: to 275%; P < 0.02). Suckling-induced release of oxytocin was significantly reduced using Ca(2+)-free, EDTA-containing (10(-4) M) microdialysis fluid and further stimulated by high K(+)- (56 mM), veratridine-containing (50 microM) microdialysis fluid. The opioid antagonist naloxone whether given by subcutaneous injection (5 mg/kg) or directly into the supraoptic nucleus by microdialysis (5 x 10(-6) M) or microinjection (1.5 microliters, 10(-6) M) did not further enhance oxytocin release within either the supraoptic or paraventricular nuclei during parturition. In contrast to the selective release of oxytocin within the supraoptic and paraventricular nuclei during parturition and suckling, direct osmotic stimulation of the nuclei by microdialysing hypertonic medium (artificial cerebrospinal fluid; 1 M NaCl) increased intranuclear release of both oxytocin and vasopressin which was further enhanced after replacement of hypertonic with isotonic fluid. This rebound phenomenon served to confirm the precise location of the microdialysis probe ante mortem and the ability of the nuclei to adequately respond to the osmotic stimulus at the end of the experiment. The study has shown that oxytocin is released in the supraoptic and paraventricular nuclei during parturition as well as in lactation unrestrained by endogenous opioids during parturition. This intranuclear release of oxytocin may act by local positive feedback stimulation of oxytocin neurons to excite further oxytocin release in the brain and into blood during both parturition and lactation.

Simultaneous Microdialysis in Blood and Brain: Oxytocin and Vasopressin Release in Response to Central and Peripheral Osmotic Stimulation and Suckling in the Rat

Simultaneous microdialysis in blood and brain has been used to monitor the release of both oxytocin and vasopressin into the systemic circulation (jugular vein/right atrium) and within the hypothalamic supraoptic nucleus of rats. Both home-made probes for blood and brain microdialysis revealed detectable nonapeptide concentrations under basal conditions and differential responses to a variety of stimuli. In urethane-anesthetized male rats, bilateral stimulation of the supraoptic nucleus by microdialyzing hypertonic medium (1 M NaCl) not only significantly increased the intranuclear release of both oxytocin and vasopressin (p < 0.05), but also their release from the neurohypophysis into blood (p < 0.05). In poststimulation microdialysates sampled from blood, the nonapeptides reached basal levels again, whereas intranuclear levels were further elevated. Intraperitoneal injection of hypertonic saline, on the other hand, resulted not only in the well-known increased peripheral release of oxytocin and vasopressin (p < 0.01 each), but also in a delayed increase in intranuclear oxytocin (p < 0.05). In contrast, intranuclear vasopressin release failed to change within the 90-min period following osmotic stimulation. In conscious lactating rats, suckling increased oxytocin contents in microdialysates sampled simultaneously in blood and the supraoptic nucleus (p < 0.05 each) further validating the microdialysis techniques used. The in vivo recovery in blood of approximately 65% determined using both radiolabeled and endogenous oxytocin provides a rough estimate to assess nonapeptide concentrations in plasma from 30-min or even 10-min blood microdialysis data.(ABSTRACT TRUNCATED AT 250 WORDS)

Systemic osmotic stimulation increases vasopressin and oxytocin release within the supraoptic nucleus

Vasopressin (VP) and oxytocin (OT) are released within the hypothalamic nuclear region in response to direct microdialysis with hypertonic solutions. Experiments were performed to determine whether systemic osmotic stimulation causes changes in intranuclear peptide release within the supraoptic nucleus (SON). A hypertonic sodium chloride solution was injected intraperitoneally (ip) or intravenously (iv) and microdialysis techniques were used to simultaneously monitor central and peripheral peptide release in urethane anesthetized rats. Systemic osmotic stimuli elicited increases in intranuclear peptide release which were delayed and long‐lasting, occurring over a 2.5 h period. In contrast, plasma peptide levels peaked at 30‐min after the stimulus. The results demonstrate that increased plasma sodium elicits an increase in VP and OT release into the extracellular space of the hypothalamic SON. The different patterns of peptide release in plasma and brain point toward the possibility of independently regulated release into the different compartments.

Septal and Hippocampal Release of Vasopressin and Oxytocin during Late Pregnancy and Parturition in the Rat

The push-pull perfusion technique, in conjunction with specific radioimmunoassays, was used to monitor the release of both arginine vasopressin (AVP) and oxytocin (OXT) within distinct limbic brain areas of conscious female rats. In pregnant rats near term, the release of AVP was greater than that of virgin rats in both the ventral (p less than 0.001) and mediolateral (p less than 0.001) septal areas; similarly, release of OXT increased in the ventral septal area (p less than 0.01) at this time. In contrast, no changes in the levels of either peptide occurred in the dorsal hippocampus. In parturient rats, AVP release tended to decrease in the septal areas but increased fivefold in the dorsal hippocampus (p less than 0.001) compared to pregnant animals. In contrast, OXT levels assayed in the same perfusates did not differ from those observed in pregnant animals. Plasma levels of AVP in pregnant rats (p less than 0.05) and of OXT in parturient animals (p less than 0.01) were found to be increased over levels in virgin rats. The regionally different and peptide-specific changes in release pattern of AVP and OXT in virgin, pregnant and parturient rats may be of physiological significance in antipyresis and behaviors accompanying parturition.

Interleukin-1β Stimulates both Central and Peripheral Release of Vasopressin and Oxytocin in the Rat

Simultaneous microdialysis in the brain and blood was used to monitor the release of vasopressin and oxytocin within the hypothalamic supraoptic (SON) and paraventricular (PVN) nuclei and into the systemic circulation of urethane‐anaesthetized male rats before and after central administration of interleukin‐1β (IL‐1β). Following intracerebroventricular infusion of the cytokine (200 ng/5 μl), the content of vasopressin (up to 278% compared to vehicle‐treated control, P < 0.01 compared to vehicle‐treated control and preinfusion baseline) but not oxytocin (up to 148%, not significant) in 30‐min blood microdialysates was found to be increased. This peripheral release was accompanied by a transient rise in vasopressin (up to 163%, P < 0.05) and oxytocin (up to 182%, P < 0.05) release within the SON, the peak typically occurring during the first and second 30‐min collection intervals after IL‐1 β respectively. In contrast, in the simultaneously microdialysed PVN, both vasopressin and oxytocin failed to respond to intracerebroventricular IL‐1 β. In another series of experiments, IL‐1 β was directly infused (20 ng0.5 μl) into either the SON or PVN during microdialysis of the corresponding nucleus. The cytokine caused a significant and immediate rise in intra‐SON release of both vasopressin (up to 225%, P < 0.01) and oxytocin (up to 178%, P < 0.05). Again, in the PVN, nonapeptide release, although tending to be stimulated in response to intranuclear IL‐1 β, failed to reach statistical significance. The cytokine‐induced central and peripheral release pattern appeared to be independent of the rise in body temperature observed after IL‐1 β administration. In a third series of experiments, bilateral administration of IL‐1 β into the SON (20 ng/0.5 μl) failed to alter peripheral release of both vasopressin and oxytocin into the systemic circulation. The increase in central nonapeptide release in response to IL‐1 β shown in this paper supports the hypothesis that at least vasopressin might act to oppose central effects of the cytokine, including those on thermoregulation and behaviour, in this way contributing to the neuroendocrine‐immune dialogue at brain level.

Push‐pull Perfusion and Microdialysis Studies of Central Oxytocin and Vasopressin Release in Freely Moving Rats during Pregnancy, Parturition, and Lactation

Oxytocin (0T)-containing neurons are localized predominantly in the compact magnocellular and parvocellular divisions of the paraventricular nucleus (PVN) and in the supraoptic nucleus (SON). Unlike arginine vasopressin (AVP) neurons, OT-producing cell bodies appear to be absent in extrahypothalamic brain areas. Magnocellular hypothalamic neurons project to the neurohypophysis. Following peripheral release, OT is primarily associated with smooth muscle contraction of the female reproductive system, and AVP with water reabsorption by the kidney, vasoconstriction in certain vascular beds, and glycogenolysis in the liver. Additionally, parvocellular neurons of the PVN (OT, AVP) and of extrahypothalamic sources (AVP) project to central targets, especially limbic areasZ known to be involved in learning, memory, and behavioral performance. Since the transport of endogenous nonapeptides across the blood-brain barrier is severely re~tricted,~ central effects should be the result of centrally released rather than peripherally circulating OT and AVP, respectively. Such central effects include defervescence (AVP)4 and selective behavioral regulation (OT, AVP).5,6 The fact that different cell populations project peripherally to the neurohypophysis and centrally to limbic brain targets implies that a stimulus which provokes a release of OT and AVP into the bloodstream does not necessarily provoke a release within the

Naloxone increases the release of oxytocin, but not vasopressin, within limbic brain areas of conscious parturient rats: a push-pull perfusion study.

The influence of naloxone on the release in limbic brain areas of both oxytocin (OXT) and vasopressin, measured by radioimmunoassay, was studied in conscious parturient rats. Three consecutive 30-min push-pull perfusions (20 microliters artificial CSF/min) were made, via previously implanted guide cannulae, within the medio-lateral septum and dorsal hippocampus of parturient animals given saline or naloxone hydrochloride (5 mg/kg body weight) after delivery of the second pup. OXT release in the hippocampus, but not in the septum, was increased during parturition, compared to day 1 post partum. During the first 30-min collection period following naloxone administration, release of OXT was significantly elevated within the septum (44% compared to saline controls, p less than 0.002), but not in the dorsal hippocampus; vasopressin release was not affected. In contrast, on day 1 post partum, naloxone, administered 5 min after starting two consecutive perfusions failed to alter OXT release in septum or hippocampus in conscious rats. Naloxone, known to increase the release of OXT also from the posterior pituitary during parturition, speeded the parturition process significantly between the birth of pups 4 and 8 during push-pull perfusion of septum or hippocampus. The data suggest that endogenous opioid inhibition is involved in the regulation of central OXT release, but not vasopressin release, during parturition. Together with previous studies on OXT release from the posterior pituitary, it seems that during parturition there is coordinated endogenous opioid action on the release of OXT both into blood and into the brain.

Chapter 8: Endogenous opioids regulate intracerebral oxytocin release during parturition in a region-specific manner

Publisher Summary During parturition, the nonapeptide oxytocin (OXT) plays an important role in promoting labor. It is established that the release of OXT from the posterior pituitary into blood during the parturition process is under the inhibitory influence of endogenous opioids. OXT and the related nonapeptide arginine vasopressin (AVP) are released intracerebrally within limbic targets and hypothalamic sites of origin in response to a variety of stimuli. OXT and AVP fibres projecting for example to septal or hippocampal targets, and different structures within the hypothalamic supraoptic nucleus (SON), provide the morphological basis for central nonapeptide release. This chapter discusses if: (1) the intracerebral release of OXT and AVP by neurones projecting to the medio-lateral septum and dorsal hippocampus is inhibited by endogenous opioids during parturition, and also whether opioids act at the level of oxytocinergic structures within the target region, (2) OXT release within the SON during parturition is regulated by endogenous opioids, and (3) peripheral and septal administration of NLX, respectively, hasten the parturition process during push–pull perfusion.

Crosstalk in the magnocellular system during osmotic stimulation of one supraoptic nucleus.

Neural connections linking the four magnocellular nuclei, i.e., the paired supraoptic (SON) and paraventricular (PVN) nuclei, may contribute to the simultaneous and parallel changes in firing patterns of oxytocinergic neurons during reflex milk ejection. To investigate these neural connections in the absence of suckling, intranuclear release of oxytocin (OT) was stimulated by microdialysis of hypertonic CSF containing 1 M NaCl (HS-CSF) into the right SON area and glucose metabolism of both SONs and PVNs and the neural lobe of virgin and lactating (10-12 day) rats was mapped by the autoradiographic [14C]deoxyglucose (DG) method. OT in the microdialysates and in plasma, obtained before and after 80-90 min of dialysis with CSF or HS-CSF, was quantified by RIA. In both virgin and lactating rats, microdialysis of HS-CSF unilaterally into the SON area significantly (p < 0.05) increased release of OT in the nucleus and into plasma, which was associated with enhanced (p < 0.05) metabolic activity in the ipsilateral and contralateral SON and the neural lobe but not in either PVN. Compared with virgins, lactating rats were less active, had lower (p < 0.05) glucose utilization in the hypothalamo-neurohypophysial system, and less (p < 0.05) OT in plasma during microdialysis of HS-CSF into the SON area. The osmotic stimulus did not activate neural structures (suprachiasmatic and medial amygdaloid nuclei) near the SON in either hemisphere. Thus, neural connections or, less likely, transport of OT via the subarachnoid space, may function to recruit activation of cells in the contralateral SON following hypertonic stimulation of cells in the other SON.