Chronic Stimulation of Leptin on Food Intake and Body Weight after Microinjection into the Ventromedial Hypothalamus of Conscious Rats

A very low dose of leptin (50 ng) was microinjected into the ventro-medial hypothalamus (VMH) of each rat daily once for three days. Food intake and body weight were measured after leptin injections. Microinjection of leptin into the VMH reduced food intake by 33.3 % significantly (P<0.01) during three days of leptin injection compared to the control. Body weight was measured after 24 h, 48 h and 72 h of leptin injection. After 24 h (P<0.01) and 48 h (P<0.05) of leptin injection, body weight was reduced significantly compared to that of rat before injection. Similarly, after 72 h of leptin injection, a significant reduced body weight was observed (P<0.1). A significant (P<0.001) reduced changes of body weight were found after 24 h, 48 h and 72 h after injection into the VMH when compared to the respective controls injected with saline. The results suggest that leptin has dramatic effect on reducing body weight by inhibition of food intake. TAJ 2006; 19(2): 70-75 1 Assistant Professor, Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi-6205, Bangladesh. 2 Professor, Department of Medical Biochemistry, Ehime University School of Medicine, Ehime 791-02, Japan. Introduction The hypothalamus is involved in regulation of obesity, caused by hyperphagia and decreased energy expenditure and motility. Among the hypothalamic structures, the VMH has been recognized as the satiety center. The sympathetic output from the VMH neurons innervates the peripheral tissues and regulates metabolic function. The destruction of the VMH produces severe hyperphagia and obesity (12) showing the VMH as the key center for regulation of body weight and food intake. Some of the peptides found in the hypothalamus and in other peripheral tissues. These peptides regulate hyperphagia and other syndromes of obesity. NPY was found in the arcuate nucleus and is believed to enhance feeding behavior of animal (18). The function of leptin in feeding is opposite (19). It is speculated that leptin inhibits NPY synthesis in the brain and inhibit feeding. Therefore, the mechanism of developing obesity and its prevention is not well defined. Therefore, the present study has been undertaken to find the role of leptin in feeding and body weight and whether the VMH is involved in this phenomena. This study was also undertaken to evaluate the possible role of the VMH region as a target for leptin’s satiety signal. For this purpose, we microinjected very low doses of recombinant human leptin directly into the VMH to show the role of leptin in feeding behavior and body weight regulation. Materials and Methods Male Sprague-Daweley rats weighing 200 g to 250 g were used. They were housed in plastic cages at 25±1 C with a 12h light-dark cycle (lights on at 06.00-18.00 h) and given free access to laboratory chow and water. The rats were anesthetized with sodiumpentobarbital (40 mg per kg body weight). They were placed in steriotaxic instruments and implanted double-walled cannula to the VMH of rats according to the coordinates (10). For the VMH, the stereotaxic coordinates used were AP 5.8 (5.8 mm anterior to the interaural line), L 0.5 (0.5 mm lateral to the sagital suture), and H 9.5 (9.5 mm below the surface of the skull). They were maintained in the cages with free access of water and ad libitum. When the body weight is recovered, the rats were used for experiment. The rats were repeatedly handled during the 5to 7day recovery period to habituate them to the injection procedure. 50 ng of leptin (recombinant human leptin) dissolved in 0.5 μl saline was given to the VMH of each rat. Control rats were given 0.5 μl saline only. Rats were injected with a single dose of leptin into the VMH once for three days daily at 3 P.M. Body weight was measured in the animal laboratory with balance (Yamato). Initial food given to each rat was weighed. After three days of leptin injection, the final food was weighed. Net food intake by each rat was calculated as the difference between the initial and final food weight. Body weight was measured for 24 h, 48 h and 72 h after leptin injection. All data, expressed as means ± SE, were analyzed using analysis of variance (pared t-test) by Stat View soft ware.