Prokineticin 2 is a hypothalamic neuropeptide that potently inhibits food intake

Abstract

Objective: Prokineticin 2 (PK2) is a hypothalamic neuropeptide expressed in central nervous system areas known to be involved in food intake. We therefore hypothesized that PK2 plays a role in energy homeostasis.

Research design and methods: We investigated the effect of nutritional status on hypothalamic PK2 expression and effects of PK2 on the regulation of food intake by intracerebroventricular (ICV) injection of PK2 and anti-PK2 antibody. Subsequently, we investigated the potential mechanism of action by determining sites of neuronal activation after ICV injection of PK2, the hypothalamic site of action of PK2, and interaction between PK2 and other hypothalamic neuropeptides regulating energy homeostasis. To investigate PK2's potential as a therapeutic target, we investigated the effect of chronic administration in lean and obese mice.

Results: Hypothalamic PK2 expression was reduced by fasting. ICV administration of PK2 to rats potently inhibited food intake, whereas anti-PK2 antibody increased food intake, suggesting that PK2 is an anorectic neuropeptide. ICV administration of PK2 increased c-fos expression in proopiomelanocortin neurons of the arcuate nucleus (ARC) of the hypothalamus. In keeping with this, PK2 administration into the ARC reduced food intake and PK2 increased the release of alpha-melanocyte-stimulating hormone (alpha-MSH) from ex vivo hypothalamic explants. In addition, ICV coadministration of the alpha-MSH antagonist agouti-related peptide blocked the anorexigenic effects of PK2. Chronic peripheral administration of PK2 reduced food and body weight in lean and obese mice.

Conclusions: This is the first report showing that PK2 has a role in appetite regulation and its anorectic effect is mediated partly via the melanocortin system.

FIG. 1.

Fasting reduces hypothalamic expression of PK2. Hypothalamic expression of PK2 mRNA among ad libitum–fed, 12-h fasted, and 24-h fasted rats (n = 24 per group) is shown. Results are expressed as mean ± SEM. ***P < 0.001 versus fed group.

FIG. 2.

PK2 potently reduces food intake independent of changes in locomotor activity or energy expenditure. Food intake: Effect on 0–1 h food intake in ad libitum–fed rats (n = 10–12 per group) after ICV administration at the beginning of the dark phase of PK2 at doses of 0.005, 0.015, 0.05, and 0.15 nmol/rat (A). Effect on 0–1 h food intake in ad libitum–fed rats (n = 10–12 per group) after ICV administration at the beginning of the dark phase of PK2 at doses of 0.15, 0.50, and 1.5 nmol/rat (B). Rats (n = 10–12 per group) fasted for 24 h were injected intracerebroventricularly in the early light phase (C and D) with PK2 at doses of 0.15, 1.5, or 4.5 nmol/rat. Food intake in the first hour (C) and cumulative food intake for 24 h after injection are shown (D). Results are expressed as mean ± SEM (n = 10–12 per group). *P < 0.05, **P < 0.01, ***P < 0.001 versus saline. Locomotor activity: Effect of ICV injection of saline or PK2 1.5 nmol/rat at the beginning of the dark phase on horizontal (E) and rearing (F) movement, respectively. Data are shown as mean ± SEM for each 30-min time period (n = 10–12 per group). Horizontal black bar under the x-axis indicates dark phase and open bar indicates light phase. Energy expenditure: Effect of ICV injection of saline or PK2 1.5 nmol/rat on oxygen consumption (G). Horizontal black bar under the x-axis indicates dark phase and open bar indicates light phase.

FIG. 3.

Immunoblockade of endogenous PK2 increases food intake. The effect on 2–4 h food intake of ICV administration of control IgG or anti-PK2 antibody (10 or 30 pmol) to satiated rats (n = 10–12 per group) at the beginning of the light phase. Results are expressed as mean ± SEM. *P < 0.05 versus 30 pmol control IgG group.

FIG. 4.

PK2 mediates its effects via specific hypothalamic nuclei. A: Graphical representation of c-fos activation in hypothalamic nuclei of rats after administration of saline or PK2 (1.5 nmol/rat) into the lateral ventricle. Open bars represent saline-injected animals; filled gray bars, PK2-injected animals. Data are shown as median and interquartile range. SON, supraoptic nucleus; ARC, arcuate nucleus; PVN, paraventricular nucleus; AHA, anterior hypothalamic area; SCN, suprachiasmatic nucleus; VMH, ventromedial hypothalamus; LHA, lateral hypothalamic area; DMN, dorsomedial nucleus. *P < 0.05 versus saline. B–F: Representative brain sections showing c-fos expression in the SON (B), ARC (C), PVN (D), and AHA (E and F) of rats injected into the lateral ventricle with saline or PK2 (1.5 nmol/rat). Scale bar, 100 μm. Brain sections from rats injected with saline are shown in the panels on the left and those from rats injected with PK2, in the panels on the right. Representative brain sections showing c-fos expression in the VMH, DMN, SCN, and LHA are shown in supplementary Fig. 1. G: Effects on food intake of saline or PK2 (0.025 nmol/rat) injection into specific hypothalamic nuclei at the beginning of the dark phase into rats. Food intake consumed in the first hour after PK2 injection (black bar) is shown as mean ± SEM as a percentage of food intake consumed in the first hour after saline injection (white bar) for each nucleus. *P < 0.05, **P < 0.01 versus saline.

FIG. 5.

PK2 mediates part of its anorectic effects via the melanocortin system. A: Effect of PK2 on α-MSH release from hypothalamic explants. Peptide release is expressed as percentage of basal (n = 9–12 per treatment). *P < 0.05 versus basal. B: Effects of melanocortin receptor antagonism on anorectic effects of PK2. Food intake in the 0–2 h after injection is shown. Results are expressed as mean ± SEM. *P < 0.05 versus saline. C: Effect of ICV administration of PK2 on c-fos expression in arcuate nucleus POMC neurons. Arcuate nucleus sections from animals injected with saline (1 and 3) or PK2 (2 and 4) are shown. The green arrows indicate cells expressing only POMC mRNA; the red arrows indicate cells expressing only c-fos; dual-labeled cells are indicated by a blue arrow. 3V, third cerebral ventricle; scale bars, 100 μm (1 and 2 are shown at ×10 magnification; 3 and 4 are shown at ×20 magnification). (A high-quality digital representation of this figure is available in the online issue.)

FIG. 6.

Peripheral administration of PK2 acutely reduces food intake. A: The effect on food intake of intraperitoneal administration of saline or PK2 at doses of 2.3, 7, or 20 nmol/kg (n = 10–12 per group) in rats at the beginning of the dark phase. Food intake in the first hour after injection is shown. Results are expressed as mean ± SEM. *P < 0.05 versus saline. B and C: Effect on food intake of intraperitoneal administration of saline or PK2 at doses of 7, 20, 60, 180, or 540 nmol/kg. Food intake in the first hour after injection (B) and cumulative food intake over 24 h (C) is shown. Results are expressed as mean ± SEM. **P < 0.01, ***P < 0.001 versus saline.

FIG. 7.

Chronic peripheral administration of PK2 decreases body weight in lean and obese mice. A and B: Cumulative food intake (A) and change in body weight (B) of C57BL/6 mice (n = 10 per group) intraperitoneally injected twice daily for 5 days with either saline or PK2 (180 nmol/kg). C and D: Effects of intraperitoneal injection of saline or PK2 (540 nmol/kg per injection) twice daily for 5 days to C57BL/6 DIO mice. Cumulative food intake of the mice injected with saline or PK2 throughout the study is shown in C. The food intake of the pair-fed group was restricted to the median food intake consumed by the PK2-treated mice over the previous 24-h period. Change in body weight of the saline-treated, pair-fed, and PK2-treated mice throughout the study is shown in D. Results are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 versus saline.