ABSTRACT
The hypothalamus plays an important role in food consumption, receiving information about energy balance via hormonal, metabolic, and neural inputs. Its neurons produce neuropeptides influencing energy balance. Especially important to regulation of food consumption are certain hypothalamic structures, including the arcuate (ARC) and ventromedial (VMN) nuclei and the lateral hypothalamic area (LHA). We determined the impact of cholecystokinin (CCK) and cocaine and amphetamine regulated transcript (CART) peptide, on activity of ARC and VMN neurons and hypocretin (Hcrt) synthesizing neurons in LHA. ARC is an integrative nucleus regulating food consumption, VMN is considered to be a satiety centre, and LHA a hunger sensing centre. After overnight fasting, male C57Bl/6 mice received intraperitoneal injection of (i.p.) saline (SAL) or CCK (4microg/kg) or intracerebroventricular injection of (i.c.v.) CART peptide (0.1microg/mice) or CCK (i.p.) followed by CART peptide (i.c.v.) 5min later. Sixty minutes later, the presence of Fos or Fos/Hcrt immunostaining indicated activity of ARC and VMN neurons, as well as of Hcrt cells in LHA. CCK alone did not influence neuronal activity in any of the nuclei studied. CART peptide stimulated neurons in ARC and VMN (p<0.01) but decreased Hcrt neuronal activity in LHA (p<0.05). Co-administration of both peptides synergistically stimulated ARC neurons (p<0.01) and asynergistically inhibited LHA Hcrt neurons (p<0.01). Results indicate that CCK may modify the effect of CART peptide and thus substantially influence activity of neurons in hypothalamic structures involved in regulation of food intake.
Subject(s)
Cholecystokinin/pharmacology , Cocaine/pharmacology , Eating/drug effects , Hypothalamus , Nerve Tissue Proteins/pharmacology , Neurons/drug effects , Oligopeptides/pharmacology , Pyrrolidonecarboxylic Acid/analogs & derivatives , Animals , Appetite Depressants/pharmacology , Dopamine Uptake Inhibitors/pharmacology , Hypothalamus/anatomy & histology , Hypothalamus/drug effects , Hypothalamus/physiology , Injections, Intraventricular , Intracellular Signaling Peptides and Proteins/metabolism , Male , Mice , Mice, Inbred C57BL , Neuropeptides/metabolism , Neurotransmitter Agents/metabolism , Neurotransmitter Agents/pharmacology , Oncogene Proteins v-fos/metabolism , Orexins , Pyrrolidonecarboxylic Acid/pharmacologyABSTRACT
BACKGROUND: CART (cocaine- and amphetamine-regulated transcript) peptide and cholecystokinin (CCK) are neuromodulators involved in feeding behavior. This study is based on previously found synergistic effect of leptin and CCK on food intake and our hypothesis on a co-operation of the CART peptide and CCK in food intake regulation and Fos activation in their common targets, the nucleus tractus solitarii of the brainstem (NTS), the paraventricular nucleus (PVN), and the dorsomedial nucleus (DMH) of the hypothalamus. RESULTS: In fasted C57BL/6 mice, the anorexigenic effect of CART(61-102) in the doses of 0.1 or 0.5 microg/mouse was significantly enhanced by low doses of CCK-8 of 0.4 or 4 microg/kg, while 1 mg/kg dose of CCK-A receptor antagonist devazepide blocked the effect of CART(61-102) on food intake. After simultaneous administration of 0.1 microg/mouse CART(61-102) and of 4 microg/kg of CCK-8, the number of Fos-positive neurons in NTS, PVN, and DMH was significantly higher than after administration of each particular peptide. Besides, CART(61-102) and CCK-8 showed an additive effect on inhibition of the locomotor activity of mice in an open field test. CONCLUSION: The synergistic and long-lasting effect of the CART peptide and CCK on food intake and their additive effect on Fos immunoreactivity in their common targets suggest a co-operative action of CART peptide and CCK which could be related to synergistic effect of leptin on CCK satiety.
Subject(s)
Appetite Regulation/drug effects , Nerve Tissue Proteins/pharmacology , Sincalide/pharmacology , Thinness , Animals , Appetite Regulation/physiology , Benzodiazepinones/pharmacology , Devazepide/pharmacology , Dorsomedial Hypothalamic Nucleus/drug effects , Dorsomedial Hypothalamic Nucleus/physiology , Dose-Response Relationship, Drug , Drug Synergism , Exploratory Behavior/drug effects , Exploratory Behavior/physiology , Hormone Antagonists/pharmacology , Injections, Intraperitoneal , Injections, Intraventricular , Male , Mice , Mice, Inbred C57BL , Neurons/drug effects , Neurons/metabolism , Paraventricular Hypothalamic Nucleus/drug effects , Paraventricular Hypothalamic Nucleus/physiology , Peptide Fragments/pharmacology , Phenylurea Compounds/pharmacology , Proto-Oncogene Proteins c-fos/metabolism , Receptor, Cholecystokinin A/antagonists & inhibitors , Receptor, Cholecystokinin B/antagonists & inhibitors , Solitary Nucleus/drug effects , Solitary Nucleus/physiologyABSTRACT
CART (cocaine- and amphetamine-regulated transcript) peptides have been studied for ten years. We report specific binding of 125I-CART(61-102) to the rat adrenal pheochromocytoma PC12 cell line, both intact cells and cell membranes. Saturation binding to intact plated cells resulted in Kd of 0.48+/-0.16 nM and Bmax of 2228+/-529 binding sites/cell. 125I-CART(61-102) was also bound to PC12 cells differentiated using nerve growth factor to the neuronal phenotype with non-specific binding below 20%, and Kd of 1.90+/-0.27 nM and Bmax of 11,194+/-261 binding sites/cell. In competitive binding experiments, CART(61-102), CART(55-102) and di-iodinated CART(61-102) were bound to PC12 cell membranes with Ki in low nM range; their affinity to intact non-differentiated and differentiated cells was in low 10(-8) M range. In order to prove that iodination did not eliminate the pharmacological properties of CART, we tested the biological activity of di-iodinated CART(61-102). It decreased food intake in in vivo feeding experiment on fasted mice in a dose of 1 microg/mouse to the same extent as CART(61-102) in a dose of 0.5 microg/mouse.
