Background

The POA is a central hub for body temperature homeostasis, which receives thermosensory information from the periphery and tunes the degree of sympathetic output to brown adipose tissue (BAT), cutaneous blood vessels, and heart to control the amount of heat generation and dissipation (Nakamura, 2011). We discovered that LepRb neurons in the POA are stimulated by warm ambient temperature and mediate warm-adaptive responses that include suppression of BAT thermogenesis and food intake (Yu et al., 2016). This discovery was made mainly through chemogenetic stimulation of POA LepRb neurons by virally expressing Gq-coupled DREADD, hM3Dq, in POA LepRb neurons and injecting CNO in mice. A single IP injection of CNO can stimulate target neurons up to 10 h (Krashes et al., 2011; Rezai-Zadeh et al., 2014). This long-lasting CNO effect allows researchers to modulate target neuron activity chronically with two IP injections of CNO per day.

Optogenetics and chemogenetics have revolutionized the field of neuroscience by providing tools to selectively manipulate target neuron activity with its own unique advantages and disadvantages. For studying neurons that modulate energy balance, researchers often use an indirect calorimetry system to simultaneously measure energy expenditure and food intake for an extended period of time (Rezai-Zadeh et al., 2014; Correa et al., 2015; Qualls-Creekmore et al., 2017). Chemogenetics is best suited for this type of study thanks to slow clearance of CNO from the body and no requirement of optical devices as in optogenetics. In our study, we investigated consequences of chronic stimulation of POA LepRb neurons by DREADD with the PhenoMaster indirect calorimetry system. CNO was injected at 0.3 mg/kg twice per day for six days, and energy expenditure and food intake were continuously measured every 25 min. Body weight was measured once every morning to monitor how changed energy expenditure and food intake affected body weight. In this protocol, we describe a step-by-step procedure of using the TSE PhenoMaster indirect calorimetry system to measure energy expenditure and food intake during chronic stimulation of POA LepRb neurons by DREADD in mice.