|dc.description.abstract||The key factors shaping consumption involve a feeling of hunger (determines search for calories), satiation (underpins termination of ingestive behaviour), and reward (adjusts consumption to the 'pleasantness' of food, regardless of actual energy needs). Though each of the many feeding-related neural systems affects the select facets of appetite to a different degree, most basic research (and subsequently, also clinical) studies exploring anorexigenic potential of neuroactive molecules, focus on targeting only one system. This single-system approach has been the case for one of the most potent anorexigens identified to-date, oxytocin (OT). OT is known to promote satiation and its release coincides with, e.g., excessive stomach distension and osmolality. OT’s effects on feeding reward are weaker as they are mitigated by macronutrient composition, flavour and energy density. Its potential influence on perceived hunger is unknown. Therefore, in this thesis I wished to examine whether by combining OT with a ligand that targets those facet(s) of feeding largely unaffected by OT, a synergistic effect on feeding can be achieved. Considering that opioid receptor antagonists mainly reduce reward-driven feeding, but do not promote satiety (notably, their effect on hunger remains undefined), I focused on investigating the simultaneous effects of OT receptor stimulation and opioid receptor blockade on food intake.
As the potential impact of either opioid antagonists or OT alone on hunger perception has never been examined, prior to studying simultaneous effects of the two types of molecules on feeding, I investigated whether either OT or opioid ligands individually reduce a feeling of hunger. The first set of experiments explored whether hypophagic doses of OT would alter hunger responsiveness in male rats trained to discriminate between 22 h (hunger) vs 2 h (no hunger) food deprivation in a two-lever, operant discrimination procedure. Intraperitoneal (i.p.) OT did not decrease 22-h deprivation-appropriate responding to match that following 2-h food deprivation, thus, it did not reduce the perceived level of hunger. In order to understand mechanisms behind this ineffectiveness of OT, I used c-Fos immunohistochemistry to determine whether OT activates a different subset of brain sites under 22- vs. 2-h deprivation. I found that in sated animals, OT induces c-Fos changes in a broader network of hypothalamic and brain stem sites compared to the hungry state. Finally, by employing qPCR, I found that fasted animals had lower OT receptor mRNA levels in the brain stem, a CNS “entry” region for peripheral OT, than their ad libitum-fed counterparts. It can be concluded that OT does not diminish a feeling of hunger and that, therefore, OT's anorexigenic properties are manifested once consumption has already begun which is—at least to some extent—driven by changes in brain responsiveness to OT treatment in the hungry vs. fed state.
In the second set of experiments, I explored whether opioid receptor ligands, acting primarily on reward-related drivers of consumption and not on satiation, might also expand onto the third facet of feeding, i.e., hunger. I subsequently utilized the aforementioned hunger discrimination paradigm to measure behavioural responses associated with hunger to explore whether modulating the opioid receptor network will alter the perception of energy deficit (hunger). While a hunger-inducer, neuropeptide Y (NPY), shifted hunger-associated operant responses in 2-hr-deprived (sated) animals, which is reflective of an increase in hunger, opioid receptor agonists (DAMGO, DSLET, orphanin FQ and butorphanol) did not induce such changes. In line with that, hypophagic doses of an opioid antagonist, NTX, did not reduce the hunger discriminative stimuli induced by either 22 h deprivation, or by NPY administration in 2 h food-restricted rats. I therefore conclude that the opioid system, similarly to OT, does not affect perceived hunger.
The subsequent experiments relied on the hypothesis that by combining the effect of opioid receptor blockade on diminishing feeding reward and of OT on promoting early satiation, a synergistic effect can be achieved. This hypothesis was strengthened by a recent case report involving a male with hypothalamic obesity, in which concurrent administration of OT and NTX, synergistically affected energy balance. In the third set of studies performed in adolescent rats I indeed found that, while OT reduces deprivation-induced chow intake, and NTX reduces palatable food intake, that the combination of these drugs at subthreshold (ineffective in the context of feeding) doses produces a hypophagic effect on acute food intake. Furthermore, using c-Fos immunochemistry, I found robust changes in feeding-related brain regions within the brain stem-hypothalamic network as a result of this drug combination. I conclude that administration of combined OT and NTX acutely suppresses food intake, likely by modulating activity of brain regions associated with both satiety and reward.
Importantly, the OT-NTX synergy described above pertained to adolescent animals and to short-term food intake. Thus, in the fourth set of studies, I examined both acute feeding responses and longer-term feeding and body weight changes in adult rats treated with OT-NTX. I found that in adult animals, OT-NTX did not appear to produce a significant acute effect on standard chow intake as in adolescents, but I still observed the acute hypophagic effect in palatable feeding scenarios. Furthermore, while the drug combination indeed had a cumulative effect on reducing palatable food intake with chronic administration, the beneficial effects on caloric intake and body weight were offset by compensatory elevated intake of standard chow in the remainder of the day. The results of PCR gene expression analysis across three brain regions (hypothalamus, brain stem and nucleus accumbens) showed that the OT-NTX combination produced unique mRNA level changes in brain stem-hypothalamic networks.
Overall, evidence gathered throughout this project shows that co-administration of two molecules that do not diminish hunger, but that do affect distinct facets of feeding: OT (early satiation) and NTX (eating for pleasure), generates a synergistic effect in reducing food intake. This effect can be attained with doses of drugs that individually would be too low to mitigate feeding, most likely thanks to the unique action of the combined drugs at feeding-relevant brain circuits.||