In the hypothalamus, AMPK integrates peripheral hormonal
signals to modulate both sides of the energy balance equation,
namely, feeding and energy expenditure [5]. For example, in the
case of ghrelin, an orexigenic stomach-derived hormone, activation
of AMPK modulates mitochondrial fatty acid oxidation, ceramide
synthesis in the endoplasmic reticulum, and intracellular levels of
reactive oxygen species. This ultimately regulates the expression
orexigenic neuropeptides in the arcuate nucleus of the hypothala-
mus (ARC), leading to increased appetite [5, 29–35]. On the other
hand, inhibition of AMPK in the ventromedial nucleus of the
hypothalamus (VMH) induces energy expenditure by boosting
sympathetic tone, leading to increased expression of uncoupling
protein 1 (UCP1) in brown adipose tissue (BAT) [5]. This mecha-
nism mediates the effects of thyroid hormones (THs) [36–39],
estradiol [40], leptin [41], glucagon-like peptide-1 (GLP-1) [42],
bone morphogenetic protein 8B (BMP8B) [43, 44], and nicotine
[45] on BAT thermogenesis, as well as the browning of white
adipose tissue (WAT) induced by THs [38] and BMP8B
[44]. The relevance of this interaction is noteworthy, because
some agents with known actions on metabolic homeostasis, such
as metformin [46, 47], liraglutide [42], and olanzapine [48–51],
and also natural molecules, such as resveratrol [52, 53] and flavo-
noids [54–57], exert their actions by modulating hypothalamic
AMPK. This evidence highlights the possibility that hypothalamic
AMPK might be a potential target for the treatment of obesity
[5, 58, 59].
In physiology, functional analyses are generally related to bio-
chemical changes in proteins. One of the most commonly used
techniques to analyze these changes is Western blotting, in which
proteins are separated through electrophoresis, transferred to a
solid support, and then analyzed using antibodies for specific pro-
tein detection. Thus, for assaying AMPK function (activation or
inactivation) in the hypothalamus, it is possible to use phospho-
specific antibodies that recognize the phosphorylation of AMPKα
at Thr-172.2 Materials
2.1 Administration
of Substances
- Polyethylene tube for intracerebroventricular cannulae
(BD Intradermic®). - Non-sterile polyethylene tubing (Becton Dickinson and Com-
pany, PE-20 for rats and PE-50 for mice). - Anesthesia. Xylazine hydrochloride and ketamine.
- Surgical blade stainless steel n21.
434 Patricia Seoane-Collazo and Miguel Lo ́pez