Characterisation of hypothalamic leptin resistance during pregnancy in the rat

Leptin is primarily an adipose-derived hormone that acts in the hypothalamus to regulate body fat levels by suppressing appetite and increasing metabolic rate. Pregnancy is characterised by increased food intake and fat mass to meet the metabolic demands of this physiological state. Leptin concentrations also increase during pregnancy, but this does not prevent the pregnancy-induced hyperphagia, suggesting a state of leptin resistance. The aims of this thesis were to measure hypothalamic leptin responsiveness during pregnancy and to investigate the potential mechanisms underlying pregnancy-induced leptin resistance. The satiety response to intracerebroventricular (i.c.v) leptin was measured in fasted non-pregnant (diestrous), early pregnant (day 7), and mid-pregnant (day 14) rats. Serial blood samples collected from another group of rats demonstrated that despite initial elevated plasma leptin concentrations in pregnant rats, fasting significantly decreased leptin concentrations so that pregnant and non-pregnant groups had similar, low leptin concentrations. Leptin treatment significantly reduced food intake in non-pregnant and early pregnant rats but not in mid-pregnant rats. In addition, there was no post-fasting hyperphagic response in the pregnant rats. These results indicate that pregnant rats become resistant to the satiety action of leptin. To investigate the mechanisms underlying pregnancy-induced leptin resistance, leptin-induced activation of hypothalamic leptin-target neurons was examined. Signal transducer and activator of transcription 3 (STAT3) phosphorylation was measured in non-pregnant and mid-pregnant rats following i.c.v. administration of leptin. Western blot and immunohistochemistry analysis indicated that leptin-induced STAT3 phosphorylation was significantly reduced in the ventromedial nucleus of the hypothalamus (VMH) during pregnancy. A suppression in the amount of leptin-induced STAT3 activation was observed in the arcuate nucleus during pregnancy, yet there was no overall change in the number of leptin responsive neurons compared to non-pregnant rats. This raises the possibility of a decrease in the degree of responsiveness of arcuate nucleus neurons to leptin during pregnancy. Using double-labelled immuno-histochemistry for alpha-melanocyte stimulating hormone ( a-MSH) and leptin-induced pST AT3 it was demonstrated that proopiomelanocortin (POMC) neurons remain responsive to leptin during pregnancy. In the VMH, consistent with the reduced pSTAT3, pregnancy also induced a 2-fold reduction in mRNA for the long form of the leptin receptor (Ob-Rb), the only isoform with full signal transduction capabilities. Expression of mRNA for one of the short forms of the leptin receptor (Ob-Ra) in the choroid plexus was decreased in early and late pregnancy, suggesting that reduced leptin transport into the brain may contribute to pregnancy-induced leptin resistance. CSF/plasma leptin concentration ratios did not differ between pregnant and non-pregnant rats however, suggesting unimpaired leptin transport during pregnancy. These results indicate that pregnancy is a state of hypothalamic leptin resistance and is associated with impaired activation of the leptin-induced JAK/STAT3 signalling pathway in the VMH and arcuate nucleus, and reduced expression of Ob-Rb mRNA in the VMH. This state of leptin resistance represents an important adaptation of the maternal brain allowing increased food intake and fat mass so that the maternal body can meet the metabolic demands of pregnancy and prepare for the subsequent demands of lactation