Abstract
In addition to the various effects of natriuretic peptides (NPs) on cardiovascular systems, increasing attention is being paid to the possibility that NPs induce adipose tissue browning and activate thermogenic program. We herein established a direct intracellular temperature measurement system using a fluorescent thermoprobe and investigated the thermogenic effects of A-type NP (ANP) on brown adipocytes. The thermoprobe was successfully introduced into rat brown adipocytes, and the temperature dependent change in fluorescence intensity ratio was measured using a fluorescence microscope. After one-hour incubation with ANP, the degree of the change in fluorescence intensity ratio was significantly higher in ANP-treated (P < 0.01) adipocytes compared to untreated controls. The ANP treatment increased uncoupling protein-1 (UCP1) mRNA levels, which is one of the markers of thermogenesis in adipocytes, while the intracellular ATP content was not changed, indicating mitochondrial uncoupled respiration. Intriguingly, these thermogenic actions of ANP were more prominent when brown adipocytes were incubated at 35 °C than at 37 °C. Moreover, the increase in the intracellular temperature and the expression of UCP1 induced by ANP were cancelled by p38MAPK inhibition. Taken together, this study directly demonstrated the thermogenic actions of ANP in brown adipocytes through the use of a novel method of intracellular temperature measurement.
Highlights
The various effects of natriuretic peptides (NPs) on the cardiovascular system play a key role in the pathophysiology of heart failure
We established a system to directly measure the intracellular temperature using a fluorescent thermoprobe under a fluorescence microscope. Using this novel system to directly measure the intracellular temperature, we demonstrated that A-type NP (ANP) exerts warming effects on brown adipocytes
To assess the temperature change in greater detail, we developed an approximation formula based on the calibration (Fig. 3), and estimated the values more precisely, as follows: ANP (10−9 M); +1.55 °C, ANP (10−7 M); +2.18 °C, isoproterenol (10−7 M); +1.94 °C, CL316,243 (0.5 μM); +1.98 °C
Summary
The various effects of natriuretic peptides (NPs) on the cardiovascular system play a key role in the pathophysiology of heart failure. Uncoupling protein 1 (UCP1) is expressed in brown adipose tissue (BAT), and enables mitochondrial uncoupled respiration, rather than ATP production, allowing for the dissipation of nutritional energy as heat[12,13,14]. MRNA levels of ANP and BNP in the heart, as well as the plasma BNP levels, are elevated in mice exposed to cold environments, which supports the idea that NPs, in parallel with the sympathetic input described above, could induce the activation of the adipose tissue thermogenic program in response to cold stimulus[15]. In order to investigate the thermogenic effects of NPs on BAT more directly, we measured the intracellular temperature of brown adipocytes after ANP stimulation using this newly established measurement system
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