Figure 5.

Effect of C75 on hypothalamic neuropeptide mRNA levels in ob/ob mice fasted for 26 h. ob/ob mice were fasted for 26 h. Hypothalami from six mice were dissected at this time (time 0); another group of 26-h fasted mice was either injected i.p. with vehicle (n = 3) or with C75 (30 mg/kg body weight) (n = 18). At 2 h (n = 6), 4 h (n = 6), and 6 h (n = 6) after injection, hypothalamic tissue was dissected (time: 2 h, 4 h, and 6 h). Total RNA (5 μg) extracted from paired hypothalami was mixed with a probe mixture containing [³²P]UTP-labeled cRNAs for AgRP (a), CART (b), NPY (c), and POMC (d) mRNAs, and subjected to the RNase protection assay protocol. Protected fragments were separated by urea-PAGE and exposed to an imaging plate for 24 h. Radioactivity in the gel bands was quantified, normalized to the cyclophilin signal, and expressed as percent change relative to the mRNA level of control mice. Neuropeptide mRNA levels in mice fasted for 26 h + 6 (○) were 108% (AgRP), 94% (CART), 106% (NPY), and 80% (POMC). Values are shown as means ± SEM. **, P < 0.01 vs. value of time 0.