Yuki Takayanagi

ADVANCES IN VASOPRESSIN AND OXYTOCIN: FROM GENES TO BEHAVIOUR TO DISEASE

Oxytocin has potent central effects on feeding behaviour, as well as on social and sexual behaviours, and one likely substrate for its anorectic effect is the ventromedial nucleus of the hypothalamus. This nucleus expresses a high density of oxytocin receptors, but contains very few oxytocin-containing fibres, hence it is a likely target of neurohormonal actions of oxytocin, including possibly oxytocin released from the dendrites of magnocellular oxytocin neurones. As oxytocin release from dendrites is regulated independent of electrical activity and of secretion from the neurohypophysis, exactly how this release is regulated by metabolic and reproduction-related signals remains to be established fully. Intriguingly though, it looks as though this central release of oxytocin from magnocellular neurons might be instrumental in a fundamental shift in motivational behaviour - switching behaviour from being driven by the need to find and consume food, to the need to reproduce.

Impaired Thermoregulatory Ability of Oxytocin-Deficient Mice during Cold-Exposure

We analyzed temperature homeostasis in oxytocin-deficient (Oxt−/−) mice and found that Oxt−/− mice exhibited lower body temperatures than wild-type animals when they were exposed to cold. Oxt−/− mice also showed slightly more weight gain, but there were no obvious differences in the morphology of white and brown adipose tissues as between wild-type and Oxt−/− mice. In cold-exposed conditions, oxytocin neurons containing c-Fos immunoreactivity existed in the paraventricular nucleus of the hypothalamus. These results suggest that the central oxytocin neurons constitute part of the thermoregulatory system involved in maintaining body temperature in cold environments.

Oxytocin Receptor in the Hypothalamus Is Sufficient to Rescue Normal Thermoregulatory Function in Male Oxytocin Receptor Knockout Mice

Oxytocin (OXT) and OXT receptor (OXTR) have been implicated in the regulation of energy homeostasis, but the detailed mechanism is still unclear. We recently showed late-onset obesity and impaired cold-induced thermogenesis in male OXTR knockout (Oxtr(-/-)) mice. Here we demonstrate that the OXTR in the hypothalamus has important functions in thermoregulation. Male Oxtr(-/-) mice failed to maintain their body temperatures during exposure to a cold environment. Oxtr(-/-) mice also showed decreased neuronal activation in the thermoregulatory hypothalamic region during cold exposure. Normal cold-induced thermogenesis was recovered in Oxtr(-/-) mice by restoring OXTR to the hypothalamus with an adeno-associated virus-Oxtr vector. In addition, brown adipose tissue (BAT) in Oxtr(-/-) mice contained larger lipid droplets in both 10- and 20-week-old compared with BAT from age-matched Oxtr(+/+) control mice. In BAT, the expression level of β3-adrenergic receptor at normal temperature was lower in Oxtr(-/-) mice than that in control mice. In contrast, α2A-adrenergic receptor expression level was higher in BAT from Oxtr(-/-) mice in both normal and cold temperatures. Because β3- and α2A-adrenergic receptors are known to have opposite effects on the thermoregulation, the imbalance of adrenergic receptors is suspected to affect this dysfunction in the thermoregulation. Our study is the first to demonstrate that the central OXT/OXTR system plays important roles in the regulation of body temperature homeostasis.

Vasopressin-induced contraction of uterus is mediated solely by the oxytocin receptor in mice, but not in humans

In the non-pregnant mouse myometrium, both arginine vasopressin and oxytocin induced contractions (pD(2)=8.55+/-0.13 and 9.23+/-0.09, respectively). The effect of oxytocin was the most potent, while the maximum contractions induced by these two peptides were almost of the same magnitude. Both vasopressin- and oxytocin-induced contractions were strongly inhibited by an oxytocin receptor antagonist, CL-12-42 (d(CH(2))(5)[Tyr(Me)(2),Thr(4),Tyr-NH(2)(9)]OVT), and weakly inhibited by a vasopressin V(1a) receptor antagonist, SR49059 ((2S)1-[(2R,3S)-5-chloro-3-(2-chlorophenyl)-1-(3,4-dimethoxybenzene-sulfonyl)-3-hydroxy-2,3-dihydro-1H-indole-2-carbonyl]-pyrrolidine-2-carboxamide). Similar results were obtained in the pregnant mouse myometrium. These results suggest that not only oxytocin- but also vasopressin-induced contraction is mediated by the activation of oxytocin receptors in the mouse myometrium. A reverse transcription polymerase chain reaction study failed to reveal mRNA of the vasopressin V(1a) receptor in the mouse myometrium. In contrast, in the non-pregnant human myometrium, vasopressin-induced contraction was inhibited by SR49059. Oxytocin showed no effect on the myometrium. These results suggest that there are significant differences in the functional receptors and contractile responses to vasopressin and oxytocin in the human and mouse uteri.

Paclitaxel-2′-Ethylcarbonate Prodrug Can Circumvent P-glycoprotein-mediated Cellular Efflux to Increase Drug Cytotoxicity

PurposeThe aim of the study was to investigate whether 2′-ethylcarbonate-linked paclitaxel (TAX-2′-Et) circumvents P-glycoprotein (P-gp)-mediated cellular efflux and cytotoxicity enhanced by TAX-2′-Et activation within human culture cells transfected with a rabbit liver carboxylesterase (Ra-CES) cDNA.Materials and MethodsTAX-2′-Et transport was characterized in a human colon carcinoma cell line (Caco-2) and paclitaxel (TAX)-resistant ovarian carcinoma cells (SKOV3/TAX60). Expression of P-gp, multidrug resistance protein (MRP) 2 and Ra-CES was detected by Western blotting. Cytotoxicity against Ra-CES-expressing cells and cellular amount of TAX produced were determined by MTT assay and using HPLC, respectively.ResultsUnlike rhodamine123 and TAX, TAX-2′-Et did not exhibit polarized transport in the Caco-2 cells in the absence or presence of verapamil. P-gp levels were expressed much higher in the SKOV3/TAX60 cells than in the Caco-2 cells. MRP2 protein was not detectable in the SKOV3/TAX60 cells. Uptake by the SKOV3/TAX60 cells was similar in quantity to the amount internalized by P-gp-negative SKOV3 cells. In the SKOV3/TAX60 cells, cellular uptake of TAX-2′-Et was not altered regardless of the absence or presence of verapamil. The cytotoxicity to the untransfected SKOV3 cells induced by TAX-2′-Et was significantly lower than that induced by TAX. In the Ra-CES-expressing SKOV3 line, the EC50 value of TAX (10.6xa0nM) was approximately four-fold higher than that of TAX-2′-Et (2.5xa0nM). Transfection of Ra-CES into another TAX-resistant ovarian carcinoma cells (KOC-7c) conferred a high level of TAX-2′-Et cytotoxicity via prodrug activation. The intracellular levels of TAX produced from TAX-2′-Et in the Ra-CES-positive KOC-7c cells significantly increased compared with the levels seen in exposure of the untransfected KOC-7c cells to TAX.ConclusionsTAX-2′-Et can circumvent P-gp-associated cellular efflux of TAX. TAX-2′-Et is converted into TAX by the Ra-CES, supporting its potential use as a theoretical GDEPT strategy for cancer cells expressing high levels of P-gp. The TAX-2′-Et prodrug efficiently increased the amount of intracellular TAX, which mediates tumor cell death.

Salt loading reduces hypothalamic noradrenaline release after noxious stimuli

Salt loading reduces neuroendocrine responses to stressful stimuli. Noxious stimuli facilitate noradrenaline release in the hypothalamus and, as a result, activate oxytocin neurones. Here, we examined effects of salt loading upon plasma oxytocin concentrations and noradrenaline release in the hypothalamus after footshocks. Male rats were allowed to drink 2% NaCl for 7 days. Salt loading reduced the footshock-induced increase in plasma oxytocin concentrations and noradrenaline release in the supraoptic nucleus (SON). Acute administration of hypertonic saline also attenuated the footshock-induced noradrenaline increase in the supraoptic nucleus. In contrast, salt loading did not significantly change activation of A1 catecholaminergic neurones in the medulla oblongata, as measured by expression of Fos protein. These data suggest that salt loading presynaptically suppresses noradrenaline release in the hypothalamus and oxytocin release into the blood after footshocks.