Study On The Role Of Ghrelin In Lateral Parabrachial Nucleus On Nocturnal Feeding And Energy Homeostasis In C57bl/6j Mice

Objective:（1）To observe the effect of ghrelin injection into the lateral parabrachial nucleus（LPBN）on nocturnal feeding behavior in C57BL/6J mice;（2）To observe the effect of ghrelin injection into the LPBN on nocturnal feeding behavior in GHSR^-/-mouse and C57BL/6J mice injected with[D-Lys3]-GHRP-6;（3）To observe the effect of long-term ghrelin injection into the LPBN on body weight;（4）To observe the effect of ghrelin on the firing rate of glucose-sensitive（GS）neurons in the LPBN;（5）To observe the effect of long-term ghrelin injection into the LPBN on the protein expression of the uncoupling protein-1（UCP-1）in the brown adipose tissue（BAT）;（6）To investigate the effect of ghrelin injection into the LPBN on the c-fos expression in the nucleus.Methods:（1）The effect of LPBN administration of ghrelin on mouse feeding behavior as well as the body weight of long-term ghrelin administration was monitored by the combination of catheter implantation,nucleus microinjection and feeding behavior monitoring technique.（2）We detected the firing rate of unit neuron and observed the effect of ghrelin on GS neurons in LPBN by the in-vivo electrophysiological technique.（3）We measured the protein expression level of UCP-1 in BAT by using Western Blot.（4）Immunohistochemistry was used to observe the activation of c-fos in C57BL/6J mice after LPBN administration of ghrelin.Results:（1）In the feeding experiment,300 pmol ghrelin injected into the LPBN significantly increased the cumulative food intake at 1^st,2^nd,3^rd hour compared with the 0.9%NaCl group.(1^st h 0.81±0.07 vs.0.31±0.09,P<0.05;2^nd h 1.10±0.09 vs.0.64±0.05,P<0.05;3^rd h1.40±0.13 vs.1.04±0.09,P<0.05)（2）Compared with the 0.9%NaCl injection,300 pmol ghrelin significantly increased nocturnal cumulative food intake in the first 3 hours.But the orexigenic effect could be blocked by[D-Lys3]-GHRP-6 and abolished in GHSR^-/-mice injected with ghrelin in the LPBN.（3）After long-term administration in the LPBN for 6 days,300 pmol ghrelin significantly increased the body weight gain from the 4^th day compared with the 0.9%NaCl injection.(4^thh d 1.45±0.15 vs.0.82±0.22,P<0.05;5^th d 1.97±0.26 vs.1.17±0.25,P<0.05;6^thh d 2.19±0.28vs.1.12±0.20,P<0.05)（4）We recorded 34 GS neurons in LPBN,44.1%of them were glucose-excited（GE）neurons and 35.3%of them were glucose-inhibited（GI）neurons.Ghrelin（1.5×10^-88 M）could significantly increase the firing rate of GE neurons（3.19±0.64 Hz to 6.56±1.21 Hz,P<0.05）and decrease the firing rate of GI neurons compared with the control group.（3.66±0.77 Hz to1.51±0.34 Hz,P<0.05）（5）After the long-term administration of 300 pmol ghrelin in LPBN for 6 days,there was no significance in the protein expression level of UCP-1 compared with 0.9%NaCl injection.（P>0.05）（6）Compared with the 0.9%NaCl injection,300 pmol ghrelin injected into the LPBN significantly increased c-fos expression in LPBN.（179.64±10.76 vs.129.88±14.67,P<0.05）Conclusion:Ghrelin injected into LPBN significantly increased nocturnal cumulative food intake through the interaction with GHSR in C57BL/6J mice;ghrelin mainly influenced the firing rate of GI neurons in LPBN;long term ghrelin administration increased the body weight gain;LPBN ghrelin promotes the c-fos expression in LPBN.