Computational protocol: Preoptic leptin signaling modulates energy balance independent of body temperature regulation

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Protocol publication

[…] 3-month-old Leprflox/flox mice were injected with AAV-Cre:GFP (6 males and 10 females) or AAV-GFP (6 males and nine females) in the POA. Food was switched from regular chow to high fat diet (58 kcal% fat, D12331, Research Diets) right after viral injections. Weekly body weight and food intake were measured from week −3 to week 15. Weekly food intake measurement was interrupted while mice were in TSE for indirect calorimetry at week −3, week 4 and week 10. Body composition was measured at week −3, week 1, week 4, week 7, week 10 and week 13 (Minispec LF110, Bruker). Adiposity index was calculated from fat mass divided by lean mass.While mice were in TSE, the ambient temperature was maintained at 22°C and mice were tested for adaptive responses to ambient temperature changes. For a warm-adaptation testing, the ambient temperature was changed to 30°C from CT 11 hr to 1 hr next morning (total 14 hr). For a cold-adaptation testing, the ambient temperature was lowered to 10°C from CT 1 hr to 9 hr (total 8 hr). Different times of the day were used for two temperature conditions to facilitate the detection of changes in energy expenditure and food intake.We checked the estrous cycle of female mice, starting at week 5 for 19 days by checking vaginal cytology as previously described (; ). Briefly, we flushed the vaginal cavity of female mice with sterile saline (~10 μl) 3–4 times and the final flush was collected on a glass slide. Vaginal cytology was observed unstained with a light microscope (Olympus BX51). The proestrus followed by the estrus was defined as one cycle ().On the day of tissue harvest, mice were fasted for 4 hr before leptin (5 mg/kg, i.p.) was injected 1 hr prior to tissue harvest. Brain, liver, BAT, inguinal WAT, gonadal WAT, calf muscle, and trunk blood were collected for each mouse except for one male that was found dead in week 14. Collected peripheral tissues were sliced into multiple pieces and instantly frozen with dry ice. […]

Pipeline specifications

Software tools POA, MUSCLE
Application Nucleotide sequence alignment
Organisms Mus musculus