Activators
of Hypothalamic AMPK
1.Ghrelin. ICV administration of ghrelin to fed satiated rats
induced a time-dependent increase in AMPK and ACCαphos-
phorylation levels, peaking acutely at 1 and 2 h and returning
to normal levels by 6 h posttreatment [30, 32, 63]. Rats
received an ICV administration of 5μg of ghrelin in 5μLof
saline or vehicle [30, 32, 63].
2.5-Aminoimidazole-4-carboxamide-1-d-ribofuranoside
(AICAR).In rats, ICV administration of the AMPK activator
AICAR increases AMPK phosphorylation in the hypothalamus
or restore a previously inhibited state. In mice, a single dose of
AICAR induces hypothalamic AMPK activation within 6 h. For
acute pharmacological AMPK activation with AICAR in rats,
inject a single dose of 5μg dissolved in DMSO, and changes in
pAMPK levels can be induced with treatment after 30 min to
2–4 h [40, 62]. For acute ICV treatment in mice, inject 3μgof
AICAR dissolved in saline and sacrificed after 6 h [64].Inhibitors
of Hypothalamic AMPK
1.Thyroid hormones. Either hyperthyroidism (excess of circulating
levels of thyroid hormones) or central administration of triio-
dothyronine (T3) decreases the activity of AMPK in the hypo-
thalamus [36–39]. In this sense, stereotaxic administration of
T3 into the VMH caused a rapid decreased of pAMPK in this
nucleus [36, 39, 65]. The induction of hyperthyroidism is
achieved by chronic subcutaneous (SC) administration ofL-
thyroxine (T4; 100μg/day, dissolved in 200μL of saline) or
saline for a period of 3 weeks (21 days) [36, 38, 39]. For acute
T3 ICV treatment (1 h), inject a single dose of 4 ng dissolved in
5 μL of DMSO [36]. For chronic ICV treatment, in rats and
mice, administrate a single ICV daily injection of T3 (for rats
4 ng/day for 5 days, dissolved in 5μL of saline; for mice 8 ng/
day, during 5 days, dissolved in 1μL of saline) at 8 p.m. just
before turning the lights off [36]. In the case of targeting
specific nuclei in the rat, acutely inject 8 ng of T3 in 1μLof
1:50 saline/DMSO or vehicle as control during 3–12 h
[36–39]. For chronic treatment delivered via a permanent
stainless steel cannula connected to an osmotic minipump
flow moderator, rats receive a 4 ng/day of T3 (in saline con-
taining 1 mM NaOH) or vehicle as control.
2.Estradiol. 17 β-Estradiol (E2), as the major endogenous estro-
gen, induces a decrease in the phosphorylation of hypothalamic
AMPK in female rats when given either ICVor injected stereo-
taxically in the VMH [40, 66]; nevertheless, when injected
directly into the ARC, no clear effect is detected. For the ICV
treatment, female rats are administered with a dose of 10 mM
17 β-estradiol in 5μL of volume (dissolved in DMSO/saline
1:1), every 12 h, just before the start of the photoperiod,
during 7 days [40, 66]. For the nucleus-specific injections,Methods for Brain AMPK 439