CPT1A in AgRP neurons is required for sex-dependent regulation of feeding and thirst

Abstract

Background Fatty acid metabolism in the hypothalamus has an important role in food intake, but its specific role in AgRP neurons is poorly understood. Here, we examined whether carnitinea palmitoyltransferase 1A (CPT1A), a key enzyme in mitochondrial fatty acid oxidation, affects energy balance. Methods To obtain Cpt1aKO mice and their control littermates, Cpt1a(flox/flox) mice were crossed with tamoxifen-inducible AgRPCreERT2 mice. Food intake and body weight were analyzed weekly in both males and females. At 12 weeks of age, metabolic flexibility was determined by ghrelin-induced food intake and fasting–refeeding satiety tests. Energy expenditure was analyzed by calorimetric system and thermogenic activity of brown adipose tissue. To study fluid balance the analysis of urine and water intake volumes; osmolality of urine and plasma; as well as serum levels of angiotensin and components of RAAS (renin–angiotensin–aldosterone system) were measured. At the central level, changes in AgRP neurons were determined by: (1) analyzing specific AgRP gene expression in RiboTag–Cpt1aKO mice obtained by crossing Cpt1aKO mice with RiboTag mice; (2) measuring presynaptic terminal formation in the AgRP neurons with the injection of the AAV1-EF1a-DIO-synaptophysin-GFP in the arcuate nucleus of the hypothalamus; (3) analyzing AgRP neuronal viability and spine formations by the injection AAV9-EF1a-DIO-mCherry in the arcuate nucleus of the hypothalamus; (4) analyzing in situ the specific AgRP mitochondria in the ZsGreen-Cpt1aKO obtained by breeding ZsGreen mice with Cpt1aKO mice. Two-way ANOVA analyses were performed to determine the contributions of the effect of lack of CPT1A in AgRP neurons in the sex. Results Changes in food intake were just seen in male Cpt1aKO mice while only female Cpt1aKO mice increased energy expenditure. The lack of Cpt1a in the AgRP neurons enhanced brown adipose tissue activity, mainly in females, and induced a substantial reduction in fat deposits and body weight. Strikingly, both male and female Cpt1aKO mice showed polydipsia and polyuria, with more reduced serum vasopressin levels in females and without osmolality alterations, indicating a direct involvement of Cpt1a in AgRP neurons in fluid balance. AgRP neurons from Cpt1aKO mice showed a sex-dependent gene expression pattern, reduced mitochondria and decreased presynaptic innervation to the paraventricular nucleus, without neuronal viability alterations. Conclusions Our results highlight that fatty acid metabolism and CPT1A in AgRP neurons show marked sex differences and play a relevant role in the neuronal processes necessary for the maintenance of whole-body fluid and energy balance.

Cpt1a silencing in AgRP neurons improves cognitive and physical capacity and promotes healthy aging in male mice

“…We previously observed that Cpt1a KO mice had reduced projections from AgRP neurons to the paraventricular nucleus (PVN) of the hypothalamus, enhancing the HPA axis and sympathetic tone. We also observed a decrease in blood levels of testosterone (Zagmutt et al, 2023).…”

“…In addition, the ability of aged mice lacking Cpt1a to develop the same level of exercise capacity as adult mice is remarkable because these animals do not show any features characteristic of decline due to aging (Graber et al, 2021). This may be a consequence of the reduced food intake observed in Cpt1a KO mice (Zagmutt et al, 2023), similar to that observed in mice under dietary restriction (Fahlström et al, 2012;Peters et al, 2022). Furthermore, in the last decade, AgRP neurons have been associated with neuronal circuits of nonfeeding behavior, including those related to reward, anxiety, and compulsive behavior (Dietrich et al, 2015;Miletta et al, 2020).…”

AgRP neuron activity promotes associations between sensory and nutritive signals to guide flavor preference

“…To our knowledge, this is the first report of AgRP neuron stimulation causing sexually dimorphic feeding behavior, although an interesting recent study demonstrated sex differences in how stress changes preference for AgRP neuron stimulation: a subset female, but not male, mice prefer rather than avoid AgRP neuron stimulation after stress [31]. Further, there are known sex differences regarding the function of metabolic signaling pathways in AgRP neurons [32; 33] and Agrp protein expression [34–36]. Because AgRP neuron stimulation increases food intake equally in males and females (Figure 2G), it will be interesting to investigate the mechanisms that underlie the observed sex differences specifically related to FNL.…”

DEFICIENCY OF THE NUTRIENT SENSOR CPT1c IN SF1 NEURONS DISRUPTS THE ENDOCANNABINOID SYSTEM RESULTING IN COMPROMISED SATIETY AND FUEL SELECTION UPON FAT INTAKE

“…Stereotaxic surgery to target VMH was carried out with the following coordinates: 1.5 mm posterior from Bregma, ± 0.5 mm lateral to midline and 5.8 mm deep. Purified AAVs expressing mCherry under Cre activity (AAV-CRE-mCherry) (Zagmutt et al, 2023) were injected bilaterally over 10 minutes through a 33-gauge injector connected to a 5 µL Hamilton® Syringe (65460-02, Hamilton Company, Reno, USA) and an infusion pump. 0,4 nL with a viral titer of 1,23x10 12 pfu/mL were infused in each injection site.…”

“…To quantify the adipocyte area, three representative images from each adipose tissue section were taken with a 20x objective using a high-sensitivity camera (Leica MC 190 HD Camera) and microscope. Images were analyzed with the Adiposoft software as described in Zagmutt et al, 2023. Images were analyzed and adipocytes were highlighted if they met the following criteria:…”