The effect of central administration of prolactin on food intake in virgin female rats is dose-dependent, occurs in the absence of ovarian hormones and the latency to onset varies with feeding regimen

Lactation in mammals is characterized by a marked hyperphagia and significantly elevated levels of prolactin (PRL). Several recent experiments in our laboratory have provided evidence for a causal relationship between PRL and hyperphagia. The present series of studies revealed that PRL injected intracerebroventricularly (i.c.v.) twice daily for ten days to free feeding virgin female rats produced a dose-dependent increase in food intake without disrupting vaginal cyclicity, that the hyperphagic effect of centrally administered PRL was not dependent on the presence of ovarian hormones, that it did not selectively potentiate feeding during the light or dark phase of the daily light cycle and that the latency of the feeding response to i.c.v. PRL administration was shorter in food restricted animals. Taken together, these results support the idea that PRL acts centrally to potentiate food intake.     

In vivo release and gene upregulation of brain prolactin in response to physiological stimuli

Although prolactin (PRL) actions and expression in the brain have been shown, dynamic changes in its intracerebral release and gene expression have still not been demonstrated. Using push-pull perfusion, the in vivo release of PRL was monitored within the paraventricular nucleus (PVN) and medial preoptic area (MPOA) of virgin female, lactating and male rats in response to various stimuli. Perfusion with a depolarizing medium (56 mm K(+)) increased local release of PRL within both the PVN (P < 0.05) and MPOA (P < 0.05) of urethane-anaesthetized rats, indicating release from excitable neuronal structures. The PRL in perfusates was verified by radioimmunoassay, Nb2 cell bioassays and western blot. Systemic osmotic stimulation (3 m NaCl i.p., 8 mL/kg b.w.) raised PRL concentration in plasma (P < 0.01) but not within the PVN, suggesting independent release from the pituitary and in distinct brain regions. Immobilization for 30 min increased PRL release within the PVN (P < 0.05) and the MPOA (P < 0.01) of virgin female and male (P < 0.05 each) rats and increased hypothalamic PRL mRNA expression (P = 0.008) after 30 and 90 min as revealed by real-time polymerase chain reaction. This indicates a stress-induced activation of both PRL release from and synthesis in hypothalamic neurons. Additionally, PRL was significantly released within, but not outside, the PVN (P < 0.01) and the MPOA (P < 0.05) of lactating rats during suckling and this was accompanied by a significant increase of PRL mRNA (P < 0.05) in the hypothalamus 60 min after suckling. This is the first demonstration of stimulus-induced, locally restricted release and gene upregulation of PRL within the brain, emphasizing the involvement of this 'novel' neuropeptide in various brain functions.     

Central implants of diluted estradiol: independent effects on ingestive and reproductive behaviors of ovariectomized rats

The present experiment was undertaken to evaluate the hypothesis that the effects of estrogens on feeding and sexual behaviors are organized separately within the brain. Thirty-three ovariectomized rats were implanted with bilateral guide cannulae aimed at either the paraventricular nucleus (PVN), medial preoptic area (MPOA), or posterior hypothalamus (PH). Subjects that received PVN implants were stimulated with either undiluted estradiol, a 3:1, or 10:1 mixture of cholesterol and estradiol. Animals in the other groups were treated with undiluted estradiol. All females were stimulated unilaterally with cholesterol and estradiol, yielding a total of 66 stimulation sites. Histological analysis revealed that, compared to cholesterol implants, undiluted estradiol in the PVN reduced food intake and body weight. More importantly, diluted estradiol implants in the PVN significantly lowered food intake and body weight. In contrast, undiluted estradiol in the MPOA, PH, or ventromedial hypothalamus (VMH) had no significant effects on feeding or body weight. Analyses of variance revealed significant main effects of implant location on female sexual behavior. Newman-Keuls tests indicated that diluted estradiol implants in the PVN produced lordosis quotients and quality scores that were significantly lower than those obtained with VMH implants. The possibility that the behavioral changes observed were due to peripheral rather than central effects of the hormone was evaluated by comparing the results of implants that produced vaginal cell cornification to those that did not. There were no significant differences between these groups on any of the other dependent variables, indicating that peripheral estradiol sufficient to induce vaginal cell cornification was neither necessary nor sufficient to account for the behavioral changes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Functional mapping of neural sites mediating prolactin-induced hyperphagia in doves

Microinjections of prolactin (PRL) into the ventromedial nucleus of the hypothalamus (VMN) or the preoptic area (POA) have been previously shown to increase food intake and body weight in ring doves. In an attempt to corroborate these results and to provide a more complete map of PRL-sensitive brain sites mediating the orexigenic action of PRL, a microinjection procedure was employed in the present study that delivered PRL or saline vehicle in extremely small volumes (10 nl/injection) to a variety of diencephalic sites in dove brain that had been previously demonstrated to contain high concentrations of PRL receptors. Estimates obtained from one female subject given a single 10 nl injection of [¹²⁵I]ovine PRL into the VMN supported the claim that such injection volumes resulted in limited diffusion, as 80% of the tissue radioactivity was found within a 280 mm area surrounding the injection site at 30 min after injection. Food intake of cannulated male doves in the mapping study was monitored daily during a 6 day baseline period, an initial 4 day treatment period, a 6–12 day post-treatment recovery period, and a second 4 day treatment period. Approximately half of the birds received PRL injections (50 ng/10 nl twice daily) and saline vehicle injections (10 nl twice daily) during the first and second treatment periods, respectively, while remaining birds received these treatments in the reverse order. No significant changes in food intake across baseline, vehicle, post-treatment, or PRL treatment periods were observed in birds with injection sites in the lateral POA, paraventricular nucleus of the hypothalamus (PVN), or the medial-basal hypothalamic region between the tuberal hypothalamus (TU) and VMN. In contrast, injections of PRL into the VMN area, medial POA, or TU resulted in average daily food intake values that significantly exceeded those recorded during other periods. The most robust feeding response was seen in the VMN group, where PRL injections resulted in a 58% increase in food intake over that recorded during injection of vehicle. This increase was significantly greater than that observed following PRL injections into the mPOA (26%) or the TU (32%). These findings suggest that the VMN may be a primary site of PRL action in promoting hyperphagia in this species, although PRL effects at other diencephalic loci, such as the mPOA and TU, may also contribute to the orexigenic action of this hormone.     

Effects of PVN lesions on the responsiveness of female rats to estradiol.

Previous research has shown that the paraventricular nucleus of the hypothalamus (PVN) is an important site of action for the effects of estradiol on feeding behavior. The recent finding that estrogenic stimulation of the PVN lowers food intake without inducing lordosis suggests that the effects of estradiol on feeding and sexual behaviors are organized separately within the brain. Whether the effects of estradiol on food intake can be attenuated by PVN lesions is therefore a question of practical and theoretical interest. In this experiment we examined the behavioral responsiveness of females with PVN lesions to peripheral treatment with estradiol. 32 adult, female rats received either bilateral or sham lesions of the PVN. All subjects were ovariectomized 2 weeks after the lesion. 2 Weeks following ovariectomy, half of the animals were injected with 2 micrograms of estradiol benzoate (EB) for 3 days, and half were injected with the oil vehicle. 10 days later, the treatment conditions for each subject (oil or EB) were reversed. Histological analysis indicated that 9 females had bilateral lesions of the PVN and 4 had bilateral lesions of the dorsomedial nucleus of the hypothalamus (DMN); 11 animals received sham lesions. Compared with oil treatment, EB injections significantly lowered water intake and body weight gain in all groups. However, food intake was suppressed in the DMN and sham but not in PVN-lesioned females. In addition, statistical analyses indicated that EB treatment induced similar levels of female sexual behavior in all groups. Thus, PVN lesions did not interfere with the ability of estradiol to stimulate lordosis.(ABSTRACT TRUNCATED AT 250 WORDS)     

Chronic infusion of norepinephrine and clonidine into the hypothalamic paraventricular nucleus

Previous experiments have shown that acute injection of NE and CLON into the PVN initiates a short-term feeding response in satiated rats. This study examined, in brain-cannulated rats, the impact of remote, chronic injections of NE, CLON, or saline on daily food intake and body weight gain. Over a period of 14 days, NE was infused into the PVN, either continuously at a rate of 12 nm/microliter/hr, or discretely at a rate of 6 nm/microliter/sec. In addition, the alpha 2-adrenergic agonist CLON was infused into the PVN discretely at a rate of 3 nm/0.5 microliter/30 sec. Relative to saline infusion, chronic (continuous or discrete) stimulation of the PVN with either of these drugs was effective in potentiating daily food intake by 12-19% and in increasing body weight gain, from approximately 1.5 g/day to 3.3 g/day. This evidence indicates that medial hypothalamic NE, especially within the PVN, is sufficiently robust to alter long-term feeding patterns and body weight regulation.     

Paraventricular nucleus lesions exaggerate dietary obesity but block photoperiod-induced weight gains and suspension of estrous cyclicity in Syrian hamsters

Lesions of the paraventricular nucleus (PVN) of the hypothalamus block short photoperiod-induced testicular regression in Syrian hamsters. We examined the effects of PVN or sham lesions on the short photoperiod-induced increases in body weight and adiposity in female Syrian hamsters. PVN lesions did not affect body weight when hamsters were housed in a long photoperiod (LD, 16:8) and fed Purina laboratory rodent chow (No. 5001). However, when fed a high-fat diet both groups gained weight, and the hamsters with PVN lesions gained approximately twice as much as the sham-operated controls. When the hamsters were exposed to a short photoperiod (LD, 8:16), only the hamsters with sham lesions displayed the typical increase in body weight. No further increase in body weight or parametrial fat pad weight was seen when the hamsters with PVN lesions were exposed to the short photoperiod. The lack of a short photoperiod-induced increase in body weight gain in hamsters with PVN lesions seems unlikely to be due to a "ceiling effect" on body weight gain because we have routinely observed neurally intact, melatonin-treated female Syrian hamsters with body weight in excess of 250 g. Finally, the short photoperiod interrupted estrous cyclicity in sham-lesioned hamsters but not in those with PVN lesions. Thus, PVN lesions exaggerate dietary obesity but prevent short photoperiod-induced weight gains and vaginal acyclicity in female Syrian hamsters.     

Identification of central sites involved in butorphanol-induced feeding in rats

Butorphanol (BT), a mixed kappa- and mu-opioid receptor agonist, induces vigorous food intake in rats. Peripheral injection of BT seems to increase food intake more effectively than intracerebroventricular administration. To further elucidate the nature of BT's influence on consummatory behavior, we examined which feeding-related brain areas exhibit increased c-Fos immunoreactivity (IR) following subcutaneous injection of 4 mg/kg body weight BT, a dose known to induce a maximal orexigenic response. We also evaluated whether direct administration of BT into the forebrain regions activated by peripheral BT injection affects food intake. Peripheral BT administration induced c-Fos-IR in the hypothalamic paraventricular nucleus (PVN), central nucleus of the amygdala (CeA), and nucleus of the solitary tract (NTS). However, 0.1-30 microg BT infused into the CeA, failed to increase food intake 1, 2, and 4 h after injection. Only the highest dose of BT (30 microg) injected into the PVN increased feeding. These results suggest that the PVN, CeA, and NTS mediate the effects of peripherally-injected BT. The PVN or CeA are probably not the main target sites of immediate BT action.     

Central administration of aminoprocalcitonin inhibits food intake and stimulates the hypothalamic-pituitary-adrenal axis in rats via the corticotrophin-releasing factor system

Aminoprocalcitonin (N-PCT), a neuroendocrine peptide derived from procalcitonin, reduces food intake and body weight when administered centrally in rats. We have recently shown that N-PCT is expressed in brain areas known to be involved in energy homeostasis, including the paraventricular nucleus (PVN) of the hypothalamus, which contains a prominent population of corticotrophin-releasing factor (CRF)-synthesising neurones. CRF plays a pivotal role in the regulation of the hypothalamic-pituitary adrenal (HPA) axis and food intake. However, little is known about functional interactions of N-PCT and CRF. In the present study, we found endogenous N-PCT protein in the rat PVN. We also showed N-PCT immunoreactivity in PVN co-localised with NeuN, a neuronal marker, or glial fibrillary acidic protein, an astrocyte marker. Double staining immunohistochemistry revealed that N-PCT co-localised with CRF in parvocellular neurones of the PVN. Intracerebroventricular N-PCT administration increased CRF mRNA and content in the hypothalamus, suggesting that N-PCT stimulates the HPA axis and suppresses food intake and body weight via CRF-dependent pathways. In keeping with this, i.c.v. co-injection of D-Phe-CRF(12-41), a CRF receptor antagonist, significantly attenuated N-PCT-induced reduction in food intake and body weight in a dose-dependent manner. Furthermore, i.c.v. administration of N-PCT increased plasma adrenocorticotrophic hormone and corticosterone concentrations and induced the expression of Fos protein, a marker of neuronal activity, in parvocellular CRF neurones. These data collectively support the hypothesis that N-PCT inhibits food intake and body weight and stimulates the HPA axis via CRF-mediated pathways.     

Oestradiol Modulates the Effects of Leptin on Energy Homeostasis by Corticotrophin‐Releasing Factor Type 2 Receptor

In addition to its action in the control of the hypothalamic‐pituitary‐adrenal axis, corticotrophin‐releasing factor (CRF) has been described as an anorexigenic neuropeptide, modulating food intake and energy expenditure. CRF synthesis is influenced by leptin, which would act to increase CRF neurone activation in the paraventricular nucleus (PVN). Gonadal hormones also participate in the regulation of energy homeostasis. The reduction of food intake and body weight gain in ovariectomised (OVX) rats treated with oestradiol is associated with an increase in CRF mRNA expression in the PVN. The present study aimed to investigate the role of CRF as a mediator of leptin responsiveness in the presence of oestradiol. Wistar female rats were bilaterally OVX and divided into three groups: OVX, OVX+E (i.e. treated with oestradiol) and OVX+PF (i.e. OVX pairfed with OVX+E). The rats received daily s.c. injections of either oestradiol cypionate or vehicle for 8 days. To evaluate the role of CRF on the effects of leptin, we performed an i.c.v. leptin injection (10 μg/5 μl) with or without previous i.c.v. treatment with an CRF‐R2 antagonist. We observed that oestradiol replacement in OVX rats reduced body weight gain and food intake. The effects of exogenous leptin administration with respect to decreasing food intake and body weight, and increasing uncoupling protein‐1 expression in the brown adipose tissue and neuronal activation in the arcuate nucleus, were reversed by previous administration of a CRF‐R2 antagonist only in oestradiol‐treated OVX rats. These effects appear to be mediated by CRF‐2 receptor because the antagonist of this receptor reversed the action of oestradiol on the effects of leptin.     

Fluoxetine disrupts food intake and estrous cyclicity in Fischer female rats

Adult, regularly cycling female Fischer rats were injected daily with 10 mg/kg fluoxetine for 12-23 days. In the first experiment, body weight and vaginal smears were monitored daily. Fluoxetine treatment reduced body weight within the first 24 h of treatment. Fluoxetine treatment also elongated the estrous cycle, reduced blood levels of progesterone, and eliminated lordosis behavior. In the second experiment, body weight and food intake were examined and a pair-fed group was included to determine if fluoxetine-induced anorexia contributed to the disturbance of the estrous cycle. In pair-fed rats, effects similar to fluoxetine treatment were present. These results lead to the suggestion that fluoxetine's anorectic properties could disrupt the female's normal endocrine cyclicity and that this disruption could be relevant to the reduction in sexual behavior and motivation. However, when the duration of fluoxetine treatment was extended beyond 16 to 17 days, fluoxetine-treated female rats reinitiated vaginal cyclicity and showed evidence of normal sexual receptivity. In contrast, the estrous cycles of their pair-fed counterparts remained disrupted. Thus, restricted food intake appears to contribute to the disruption of the estrous cycle and elimination of sexual receptivity during fluoxetine treatment. However, compensatory changes in the serotonergic system that are associated with chronic fluoxetine administration may contribute to the gradual recovery of estrous cyclicity and sexual receptivity of the fluoxetine-treated animals.     

Effect of food deprivation and leptin repletion on the plasma levels of estrogen (E2) and NADPH-d reactivity in the ventromedial and arcuate nuclei of the hypothalamus in the female rats

The exact role of leptin in fasting has not been completely elucidated. To determine whether leptin can act in fasting to influence plasma estrogen levels and nitric oxide synthase reactivity in food regulating centers of the brain, we fasted female rats for 4 days and treated them i.p. with vehicle or 100 microg of recombinant mouse leptin as 1 ml on the 3rd and 4th day twice daily (10.00 and 17.00 h). Proestrus blood was collected at 10.00, 14.00, 18.00 and at 22.00 h, plasma obtained and assayed for estrogen (E2) and leptin levels. Verification of ovulation occurrence was by examining the oviduct for extruded ovum. The rat brains were removed and processed for nitric oxide synthase reactivity in the ventromedial hypothalamus (VMH) and arcuate nucleus (ARC) using NADPH-diaphorase histochemistry, a marker for neurons expressing NOS enzyme. Leptin effect on dependable variables such as food intake, water intake and body weight gain was also investigated. Four days fasting significantly decreased body weight, estrogen and postfast leptin levels, nitric oxide reactivity in the VMH and ARC nucleus and stopped ovulation in many (4 out of 5) rats fasted and given vehicle. Leptin treatment significantly increased plasma estrogen and postfast leptin levels, restored ovulation in many (4 out of 5) rats and increased nitric oxide reactivity in the VMH and ARC. Leptin significantly inhibited food intake, water intake and gain in body weight during recommenced feeding. These observations suggest that leptin could act in the pituitary-ovarian axis during fasting to improve reproductive function by partly stimulating estrogen secretion.     

Intermittent, chronic fenfluramine administration to rats repeatedly suppresses food intake despite substantial brain serotonin reductions

The mechanisms by which fenfluramine suppresses food intake and body weight have been linked to its ability to enhance transmission across serotonin synapses in brain. This drug initially lowers body weight and suppresses food intake, yet after repeated administration food intake soon returns to normal and body weight no longer decreases. Fenfluramine also causes rapid and prolonged reductions in brain serotonin concentrations, which might account for its loss of appetite suppression. This possibility has been evaluated in rats by assessing if intermittent, chronic fenfluramine administration could suppress food intake during each treatment period, and if so, whether such an effect occurs in the presence of reduced brain serotonin levels. Rats were injected once daily with 10 mg/kg D,L-fenfluramine for 5 days, and then received no injections for the next 5 days. Control rats received only vehicle injections. This 10-day sequence was repeated five more times. During each period of fenfluramine administration, daily food intake dropped markedly the first 1-2 days of treatment, but returned to pretreatment values by day 5. Daily food intake was normal or slightly above normal during non-injection periods. Body weight dropped modestly during each period of fenfluramine administration, and rose during each subsequent period when injections had ceased. Serotonin concentrations and synthesis rates in several brain regions were markedly reduced at early, middle, and late periods of the experiment. Despite the long-term reduction in brain serotonin pools produced by fenfluramine, the drug continues to reduce food intake and body weight. Several possible interpretations of these findings are considered, based on the multiple mechanisms through which this drug has been proposed to modify synaptic serotonin transmission.     

Leptin effects on the expression of type-2 CRH receptor mRNA in the ventromedial hypothalamus in the rat

The product of the ob gene, leptin, is thought to act in the hypothalamus to reduce food intake and body weight (b.w.) in rats and mice; however, the mechanisms of leptin action in the brain have not been fully elucidated. Corticotropin-releasing hormone (CRH) is a potent anorectic neuropeptide, and its type-2 receptor (CRHR-2) in the ventromedial hypothalamus (VMH) appears to play an important role in the expression of this anorectic effect. We explored here the impact of systemic leptin administration on CRH mRNA expression in the hypothalamic paraventricular nucleus (PVN) and CRHR-2 mRNA expression in the VMH in male rats, using in-situ hybridization histochemistry. The expression of CRH mRNA in the PVN and CRHR-2 mRNA in the VMH were increased at 2 h and 6 h, respectively, after a single intraperitoneal injection of leptin (1.0 mg/kg). Continuous subcutaneous infusion of leptin (1.2 mg/kg/day) via an osmotic minipump for 5 days increased the expression of CRHR-2 mRNA in the VMH, but not the expression of CRH mRNA in the PVN, compared with vehicle treatment. The rats that received the single or continuous administration of leptin showed reductions of food intake and b.w. compared with vehicle-treated rats. These results are consistent with our previous findings that the expression of CRHR-2 mRNA in the VMH is positively correlated with plasma leptin concentrations under various conditions, and highlight the importance of circulating leptin for the regulation of VMH CRHR-2 mRNA. The present results also raise the possibility that leptin reduces food intake and b.w. at least partially due to the enhancement of the anorectic effect of CRH via increased PVN CRH expression and/or VMH CRHR-2 expression.     

Selective hepatic vagotomy does not prevent compensatory feeding in response to body weight changes

The effects of selective hepatic vagotomy on compensatory adjustments of feeding in response to experimentally induced changes in body weight were investigated in rats. Repeated injections of slow-acting insulin increased food intake and body weight in vagotomized as well as in sham-vagotomized rats. When the treatment was stopped, the rats of both surgical groups displayed a compensatory hypophagia of similar magnitude and body weight returned to preinjection levels. In turn, after 7 days of restricted feeding (5 g/day) with a concomitant loss of body weight, rats became hyperphagic and body weight approached the normal level. Again, the feeding responses or body weight changes did not differ between vagotomized and sham-vagotomized rats. The results demonstrate that selective hepatic vagotomy does not disrupt the putative feedback-loop between body weight and feeding, and raise questions concerning the role of the liver in the lipostatic control of food intake.     

Effect of paraventricular nucleus lesions on body weight, food intake and insulin levels

Lesions of the paraventricular nucleus of the hypothalamus (PVN) produce obesity and hyperphagia. However, the underlying mechanism is unknown. The connections of the PVN with brainstem centers for autonomic control suggest that a change in autonomic function could mediate the PVN obesity syndrome. We examined this hypothesis in a series of 3 experiments, searching specifically for changes in insulin secretion. Rats with PVN lesions were hyperphagic and hyperinsulinemic, when obese. However, hyperinsulinemia could not be detected prior to the onset of obesity or following weight reduction. Subdiaphragmatic vagotomy reversed the PVN obesity and lowered insulin levels below those of sham-vagotomized rats. Since noradrenergic innervation of the hypothalamus is implicated in feeding, hypothalamic norepinephrine (NE) was depleted by injection of 6-hydroxydopamine into the central tegmental tract, posterior to the hypothalamus. The effects of NE depletion was compared with those of PVN lesions. Loss of hypothalamic NE resulted in hyperphagia with no increase in body weight and no change in insulin. Histological analyses indicated that the posterior PVN was the most effective lesion focus for producing disturbances in body weight and food intake. Although the results of these experiments implicate the autonomic nervous system in PVN obesity, basal hyperinsulinemia does not appear to be a primary feature of the syndrome.     

Commissural nucleus of the solitary tract lesions reduce food intake and body weight gain in rats

This study investigated the effects of an electrolytic lesion of the commissural subnucleus of the nucleus of the solitary tract (commNTS) on body weight, daily food and water intake, and plasma glucose and insulin in rats. In the first 6 days following brain surgery, commNTS lesioned rats reduced daily food intake by 80% compared to rats with sham lesions. After this period rats with lesions of commNTS started recovering food intake, but intake remained significantly reduced until the 12th day after surgery. A reduction in body weight was observed 4 days after surgery and reached a maximum on the 12th day. After this, a partial recovery of body weight was observed, but weight remained significantly reduced compared to weights of rats with sham lesions through the conclusion of the study. Food intake and body weight gain in other rats with partial lesions of the commNTS or with lesions outside the commNTS did not differ from rats with sham lesions with regard to those variables. Daily water intake and plasma glucose and insulin were not changed by the commNTS lesions. These results suggest that commNTS is involved with mechanisms that control food intake and body weight in rats.     

Behavioral effects of corticotropin-releasing factor: localization and characterization of central effects

Corticotropin-releasing factor (CRF) has potent behavioral effects when administered intracerebroventricularly to rats. CRF and its receptors are found in an uneven distribution in the brain. In an effort to localize the site of the anorectic effect of CRF, exogenous CRF or saline was injected into cannulas directed toward the paraventricular hypothalamic nucleus (PVN), lateral hypothalamus, ventromedial hypothalamus, globus pallidus, or striatum of rats. CRF decreased food intake only when injected into the PVN. In subsequent experiments PVN injections of CRF were shown to (1) increase grooming and movement; (2) not induce a conditioned taste aversion to saccharin in a single bottle test; and (3) inhibit the increase in feeding induced by injections of norepinephrine into the PVN. These results suggest that CRF induces not only anorexia, but also increased movement and grooming by action in the PVN.     

Fos expression in the brain induced by peripheral injection of CCK or leptin plus CCK in fasted lean mice

We previously reported a synergistic interaction between leptin and cholecystokinin (CCK) to reduce food intake through CCK-A receptors in lean mice fasted for 24 h. To identify the activated neuronal pathways, we investigated changes in Fos expression in brain nuclei 2 h after single or combined intraperitoneal (i.p.) injections of leptin (120 μg/kg) and sulfated CCK-8 (3.5 μg/kg) in male lean mice (C57BL/6) fasted for 24 h using immunohistochemistry for Fos, the protein product of the early gene, c-fos. Leptin did not increase Fos expression in the brain compared with vehicle-treated mice. CCK increased the numbers of Fos-positive neurons in the nucleus of the solitary tract (NTS)/area postrema (AP), central nucleus of the amygdala (CeA) and, to a smaller extent, in the paraventricular nucleus of the hypothalamus (PVN) (5.2-, 2.3- and 0.3-fold respectively). Injections of leptin–CCK further enhanced Fos expression by 40% in the PVN compared with that induced by CCK alone, but not in the other nuclei. Devazepide (a CCK-A receptor antagonist, 1 mg/kg, i.p.) prevented the increase in Fos expression induced by leptin–CCK in the PVN and by CCK alone in the PVN, CeA and NTS/AP. These results indicate that in fasted mice, i.p. injection of CCK increases Fos expression in specific brain nuclei through CCK-A receptors while leptin alone had no effect. Leptin in conjunction with CCK selectively enhanced Fos expression in the PVN. The PVN may be an important site mediating the synergistic effect of leptin–CCK to regulate food intake.     

Effects of intracisternal vs. intrahypothalamic 5,7-DHT on feeding elicited by hypothalamic infusion of NE.

A variety of evidence has led to suggestions that brain serotonin (5-HT) and norepinephrine (NE) interact within the medial hypothalamus to control food intake. To test the possibility that chronic decrements in 5-HT might enhance NE-induced feeding, adult male rats were prepared with permanently indwelling cannulae aimed at the paraventricular nucleus (PVN), then received either intracisternal (IC) or PVN injections of the 5-HT neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT) vs. its vehicle, 1% ascorbic acid. Over a 4-week period, IC-5,7-DHT rats showed no signs of enhanced daily feeding or drinking. However, in 40-min intake tests, feeding but not drinking was enhanced by injecting 20 nmol NE into the PVN commencing 2 weeks after neurotoxin treatment. Terminal monoamine assays confirmed that IC-5,7-DHT produced large (80-90%) depletions of brain regional 5-HT. A functional index of 5-HT terminal damage was also implied by the impaired short-term feeding responses IC-5,7-DHT rats showed to the systemic administration of the 5-HT1A agonist, 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) when tested between 3 and 4 weeks after IC treatment. Over a comparable 4-week period, PVN-5,7-DHT rats also showed no tendencies to overeat or overdrink on a daily basis. However, in contrast to IC-5,7-DHT rats, they also showed no differences in their feeding or drinking responses to NE injections into the PVN. This was so despite reliable depletions of 5-HT in the hypothalamus (-28%) and hippocampus (-71%). These results support earlier work showing that neither widespread nor localized hypothalamic damage to brain 5-HT neurons produce chronic overeating. However, the data suggest that phasic enhancements of PVN NE activity may trigger enhanced feeding when there is widespread damage to brain 5-HT neurons, although the PVN does not appear to be the brain site mediating this effect.