Sub-nanowatt resolution direct calorimetry for probing real-time metabolic activity of individual C. elegans worms

Abstract

Calorimetry has been widely used in metabolic studies, but direct measurements from individual small biological model organisms such as C. elegans or isolated single cells have been limited by poor sensitivity of existing techniques and difficulties in resolving very small heat outputs. Here, by careful thermal engineering, we developed a robust, highly sensitive and bio-compatible calorimetric platform that features a resolution of~270 pW-more than a 500-fold improvement over the most sensitive calorimeter previously used for measuring the metabolic heat output of C. elegans. Using this calorimeter, we demonstrate time-resolved metabolic measurements of single C. elegans worms from larval to adult stages. Further, we show that the metabolic output is significantly lower in long-lived C. elegans daf-2 mutants. These demonstrations clearly highlight the broad potential of this tool for studying the role of metabolism in disease, development and aging of small model organisms and single cells.

Cellular Thermometry Considerations for Probing Biochemical Pathways

“…Under physiological conditions, intracellular temperature changes are usually limited to sub-mK values, which are not directly measurable in vitro by intracellular probes, unless the cells are thermally isolated by multiple vacuum chambers [41], [42]. However, if the intracellular heat release can be stimulated by external agents, the intracellular temperature changes can reach measurable limits, as we show in Figure 11.…”

“…The former is more likely since it supports the reduction in the effective thermal conductivity to values below that of typical proteins (Figure 3). Typical calorimetry techniques [41], [42] for biological cells measure the temperature of the surrounding medium to estimate the total heat released from a cell. Such techniques inherently assume an effective thermal conductivity for the cell and the surrounding medium during the calibration of the thermal resistance of the calorimetry cell using an external heater.…”

“…We discussed in this section that the effective thermal conductivity is a function of the location of the heat source, the location of the measured temperature, and the local interface resistance network (Figure 4). Since the resistance to heat flow for an external heater may be different in comparison to any intracellular heat sources, previously reported calorimetry techniques [41], [42] may not be able to capture the true intracellular heat release via externally measured temperatures.…”

“…) in an isolated cell in a petri dish setting, for a range of uniform volumetric heat release inside the cell. Typically, any physiological heat release rate is < 100 nW [28], [41], [42], [62] as marked in Figure 11. To increase the expected temperatures to measurable limits (conservatively, 0.1 K), the intracellular heat release needs to be stimulated exogenously to > 10 μW.…”

“…Our model helps to understand the typically expected temperature changes and consequently the thermometry requirements for thermogenic reactions of different timescales and heat rates. Overall, our work provides a cellular heat diffusion model and the cellular thermometry requirements, which are vital for applications such as nanoparticle heating of cryopreserved tissues [35], [36], thermal ablation of tumor cells [37]- [39], thermometry-based bioenergetics studies [9], [40]- [42], etc. This paper is organized as follows.…”