Figure 3. AMPK signaling mediates deprivation- and ghrelin-induced synaptic activity.

(A) AICAR increases f_mEPSC in AGRP neurons from fed (n = 8) but not food deprived (n = 8) mice.

(B) Cpd C reduces f_mEPSC in AGRP neurons from deprived (n = 8) but not fed (n = 6) mice and blocks the ghrelin-mediated increase in f_mEPSC.

(C) AGRP neuron dialysis with the AMPK activator ZMP (3 mM) in the patch pipette internal solution (int.) does not increase f_mEPSC relative to AGRP neurons recorded with standard internal solution (nt, data from Fig. 1B).

(D) Targeted AMPK inhibition with Cpd C (int.) in AGRP neurons does not significantly (P > 0.05) change f_mEPSC after neuron dialysis (15 min).

(E) AGRP neuron firing rate is increased by bath-applied AICAR after intracellular blockade of AMPK with Cpd C (int.).

(F) AGRP neuron firing is decreased by bath-applied Cpd C after dialysis with Cpd C (int.).

(G) Targeted AMPK inhibition with Cpd C (int.) does not block ghrelin-mediated f_mEPSC increase.

(H) Ghrelin activates firing in AGRP neurons from fed (n = 8) but not deprived (n = 7) mice with postsynaptic AMPK blockade by Cpd C (int.).

(I) Glutamate receptor blockade prevents ghrelin activation of AGRP neuron firing.

(J) Paired pulse ratio (PPR) in AGRP neurons from fed and food deprived mice in 2 mM Ca²⁺.

(K) PPR in AGRP neurons from fed mice (0.5 mM external Ca²⁺) is reduced by ghrelin and this is reversed by Cpd C. Average EPSC responses from two cells are also shown (inset).

(L,M) PPR in AGRP neurons from deprived mice (0.5 mM external Ca²⁺) is unaffected by ghrelin and increased by Cpd C. (M, inset) Average EPSC response from one cell is shown.

(N) Inhibition of CAMKK with STO-609 in AGRP neurons from fed mice blocks ghrelin-mediated but not AICAR-mediated increase of f_mEPSC.

(O) AICAR does not increase f_mEPSC in the presence of ryanodine.

(P) 8-Br-cADP ribose blocks the ghrelin-mediated increase of f_mEPSC.

(Q) Diagram of the signaling pathway supported by these experiments. Pointed and “T” arrows represent activation and inhibition, respectively.

n.s. P > 0.05, *P < 0.05, **P < 0.01, ***P < 0.001. Data are represented as mean ± s.e.m. See also Figure S2.