Loss of agouti-related peptide does not significantly impact the phenotype of murine POMC deficiency

Abstract

The hypothalamic melanocortin system is unique among neuropeptide systems controlling energy homeostasis, in that both anorexigenic proopiomelanocortin (POMC)-derived and orexigenic Agouti related-peptide (AgRP)-derived ligands act at the same receptors, namely melanocortin 3 and 4 receptors (MC3/4R). AgRP clearly acts as a competitive antagonist at MC3R and MC4R but may also have an inverse agonist action at these receptors. The physiological relevance of this remains uncertain. We generated a mouse lacking both POMC and AgRP [double knockout (DKO) mouse]. Phenotyping was performed in the absence and presence of glucocorticoids, and the response to central peptide administration was studied. The phenotype of DKO mice is indistinguishable from that of mice lacking Pomc alone, with both exhibiting highly similar degrees of hyperphagia and increased body length, fat, and lean mass compared with wild-type controls. After a 24-h fast, there was no difference in the refeeding response between Pomc(-/-) and DKO mice. Similarly, corticosterone supplementation caused an equivalent increase in food intake and body weight in both genotypes. Although the central administration of [Nle⁴, d-Phe⁷]-α-MSH to DKO mice caused a decrease in food intake and an increase in brown adipose tissue Ucp1 expression, both of which could be antagonized with the coadministration of AgRP, there was no effect of AgRP alone. These data suggest AgRP acts predominantly as a melanocortin antagonist. If AgRP has significant melanocortin-independent actions, these are of insufficient magnitude in vivo to impact any of the detailed phenotypes we have measured under a wide variety of conditions.

Fig. 1.

Phenotype of Pomc^−/−/Agrp^−/− DKO mouse. A, DKO mice have a similar phenotype to Pomc null mice, displaying obesity and a lighter ventral coat color than age-matched wild-type controls. Body weight (B) and body length (C) of wild-type (WT), AgRP knockout (KO), POMC KO, and DKO male mice on normal chow. Statistical significance vs. WT mice. Lean (D) and fat mass (E) of 4- to 5-month-old male mice determined by DEXA. All data represent mean sem. ***, P < 0.001 vs. WT mice. WT (Pomc^+/+/Agrp^+/+), n = 9; AgRP KO (Pomc^+/+/Agrp^−/−), n = 8; POMC KO (Pomc^−/−/Agrp^+/+), n = 9; DKO (Pomc^−/−/Agrp^−/−), n = 11.

Fig. 2.

Food intake and energy expenditure of DKO mouse. Total (A) and lean mass (B) corrected daily food intake of 3- to 4-month-old male mice on a normal chow diet. Time course (C) and average day/night lean mass-corrected VO₂ values (D) of 3- to 4-month-old male mice determined using indirect calorimetry. Time course (E) and average day/night ambulatory movement (F) of 3- to 4-month-old male mice determined using metabolic cages. Data represent mean sem. *, P < 0.05; ***, P < 0.001 vs. WT mice. WT (Pomc^+/+/Agrp^+/+), n = 9; AgRP KO (Pomc^+/+/Agrp^−/−), n = 8; POMC KO (Pomc^−/−/Agrp^+/+), n = 9; DKO (Pomc^−/−/Agrp^−/−), n = 11.

Fig. 3.

Response of DKO mice to fasting. Cumulative food intake (A) and body weight (B) of 10- to 12-wk-old POMC KO (Pomc^−/−/Agrp^+/+) and DKO (Pomc^−/−/Agrp^−/−) female mice over 72 hours with ad libitum feeding (0–24 h), 24 h fast (24–38 h), and ad libitum refeeding (48–72 h). Data represent mean sem. POMC KO, n = 4 (white circles with dotted line); DKO, n = 7 (black squares with solid line).

Fig. 4.

Food intake and body weight after corticosterone supplementation. Cumulative food intake (A) and body weight (B) of corticosterone- (25 μg/ml drinking water) and control-treated POMC KO (Pomc^−/−) and DKO (Pomc^−/−/Agrp^−/−) female mice aged 10–16 wk. Data represent mean total cumulative food intake and mean body weight as percentage of baseline sem. *, P < 0.05, **, P < 0.01, ***, P < 0.001 vs. control-treated mice. DKO (corticosterone), n = 8 (white squares); DKO (control), n = 8 (black squares); POMC KO (corticosterone), n = 8 (white circles); POMC KO (control), n = 8 (black circles).

Fig. 5.

Central AgRP administration to DKO mice. Cumulative food intake (A), body weight (B) and quantitative RT-PCR determined expression of BAT uncoupling protein-1 (Ucp1) mRNA (C) of corticosterone-supplemented (2–5 μg/ml drinking water) DKO (Pomc^−/−/Agrp^−/−) female mice aged 10–16 wk after icv administration of 2 nmol AgRP (black circles), 2 nmol NDP-α-MSH (black squares), 2 nmol AgRP + 2 nmol NDP-α-MSH (white squares), or saline (black triangles). Data represent mean ± sem for food intake and mean body weight as percentage of baseline sem. AgRP, n = 6; NDP-α-MSH, n = 5; AgRP + NDP-α-MSH, n = 5; saline, n = 6. *, P < 0.05, **, P < 0.01, ***, P < 0.001 vs. control treatment; ¥, P < 0.01, #, P < 0.001 vs. AgRP + NDP-α-MSH treatment.