Anorexigenic effects of naltrexone in rats
The accessibility of cheap, palatable, and caloric food in the industrialized world has shifted the primary drive of eating behavior from hunger to pleasure; this change has played a significant role in the rise in the prevalence of obesity. Sugary and fatty foods are among the most preferred and, therefore, overconsumed tastants. Opioids are key neuropeptides that propel feeding for reward. Importantly, blocking the opioid receptors in laboratory animals and humans decreases intake of and preference for those diets that are highly palatable. One of those opioid receptor blockers, a non-selective antagonist naltrexone (NTX), has been used in animal and clinical studies related to excessive appetite. Consequently, NTX in combination with another molecule, bupropion, has been approved as a pharmaceutical to treat obesity. This success underscores the need for further characterization of anorexigenic potential of NTX to determine its effects in various eating behavioral contexts/scenarios, in pathophysiological conditions characterized by overeating, as well as in treatment strategies that utilize potential synergy with other molecules that decrease appetite. Therefore, as an overarching goal of this thesis I set forth to further explore anorexigenic potential of NTX as a suppressant of palatability-driven consumption. Previous studies indicate that rats cannot discriminate NTX from saline, except at very high doses. However, in those discrimination experiments, NTX-treated animals had not been fed palatable foods, i.e., diets whose consumption affects the endogenous opioid system thereby potentially leading to changes in sensitivity to opioid receptor blockade. Thus, in Specific Aim 1, I investigated the ability of rats to discriminate between NTX and saline subjected to chronic intermittent sucrose consumption. In the experiments I used the operant setting to determine whether chronic intermittent 25% sucrose solution consumption allows rats to discriminate peripherally injected NTX from saline. Subsequently, I used a feeding regimen that paralleled the discrimination training schedule and examined the consequences of peripheral NTX treatment on c-Fos immunoreactivity (IR) in feeding-related brain regions. I found that rats given intermittent access to a sucrose solution learned to discriminate between NTX doses. None of the rats given only water learned to discriminate between NTX and saline. When access to the sucrose solution was discontinued for 14 days, the rats lost this ability to discriminate. c-Fos IR analysis revealed a significant drug and diet interaction effect in various brain regions associated with feeding behavior, particularly the amygdala, nucleus accumbens, and hypothalamic sites. I conclude that unlike in animals that are maintained on a “bland” diet (thus those that do not overeat food as a result of hedonic stimulation), in individuals chronically consuming sugar, NTX evokes an interoceptive response that allows them to discriminate the drug. The heightened response of the limbic and hypothalamic areas to NTX suggests that it is associated with a broader involvement of pathways related to both homeostatic and hedonic processes. Thus far no studies have been conducted on whether NTX reduces palatability-driven feeding in pathophysiological conditions characterized by dysregulated reward processing and excessive consumption of palatable tastants. One such condition is autism spectrum disorder (ASD). Thus, in Specific Aim 2, I studied the effect of NTX on palatable food consumption and feeding-related brain activation in a valproic acid (VPA) rat model of ASD. I investigated the effect of NTX on episodic and habitual consumption of palatable food (high-fat high-sugar chow (HFHS) and sucrose solution) in VPA ASD rats compared to the control non-VPA rats. Subsequently, I assessed changes in feeding-related brain areas in response to NTX in VPA animals after two weeks of chronic intermittent sugar access using c-Fos IR. I found that, even though NTX significantly suppressed the consumption of the HFHS diet and 10% sucrose solution in VPA rats, the VPAs required a 10-times higher dose of NTX (10 mg/kg) to generate hypophagia compared to controls (1 mg/kg NTX was effective). In order to determine the difference in the neural consequences of 1 mg/kg NTX injection between VPAs and controls that parallels the lack of hypophagia after 1 mg/kg NTX, I found that c-Fos IR response to NTX in several hypothalamic areas was more pronounced in non-VPA control rats, especially those consuming sucrose. On the other hand, there was a significant drug–diet interaction for c-Fos IR in central nucleus of the amygdala (CEA) in both VPAs and non-VPA controls. It suggests that the sensitivity of feeding-related hypothalamic circuit to NTX is particularly affected by the ASD pathophysiology. Peripherally injected NTX decreases food intake. No animal studies have investigated whether NTX suppresses appetite when administered via a non-invasive intranasal (IN) route and – if so – whether this potential anorexigenic effect can be augmented by IN co-administration of another anorexigen previously shown to act synergistically with injected NTX. Thus, in Specific Aim 3, I investigated the effect of IN NTX administered either alone or with IN oxytocin (OT; peripheral OT injections decrease appetite) on palatability-driven feeding. I examined the effect of IN OT, NTX, and OT + NTX on various aspects of consumption in rats: (a) deprivation-induced standard chow intake (b) high-fat high-sugar (HFHS) chow (c) sucrose and Intralipid solutions. In this set of pilot studies, I found that - surprisingly - IN NTX and OT + NTX did not decrease intake of standard chow after deprivation and produced a minimal effect on HFHS diet, sucrose, and Intralipid consumption. When rats were exposed to habitual sucrose consumption for four weeks, neither NTX nor OT + NTX had a significant impact on the sucrose consumption. In contrast, I found that IN OT alone decreases deprivation-induced intake of standard chow as well as of HFHS chow and sucrose in nondeprived rats. In the habitual sugar consumption paradigm, acute IN OT diminishes sucrose solution intake in animals accustomed to the 2-hour/day sucrose meal regimen. Finally, I assessed c-Fos changes in response to the acute IN OT administration in rats subjected to habitual sugar consumption. I found that IN OT alters c-Fos immunoreactivity in brain areas related to energy homeostasis and reward, including the CEA, the hypothalamic paraventricular, and the arcuate nuclei. In sum, the findings of this thesis further demonstrate that NTX administered via peripheral injections in rats reduces consumption of palatable food, whereas NTX infused IN is a poor feeding suppressant. Chronic habitual consumption of palatable food facilitates the ability to discriminate peripherally injected NTX and this effect is associated with changes in neuronal activation in feeding-related brain sites. Finally, while NTX reduces palatable food consumption in VPA autistic animals, the minimum effective dose is 10 times higher than that in non-VPA controls, and this shift in sensitivity to anorexigenic effects of NTX is reflected by a reduced hypothalamic activity in NTX-treated VPAs.
Keembiya Liyanagamage, Donisha Shani Niharika
Palatability Naltrexone c-Fos immunoreactivity, Brain reward system
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