Cellular and Intravital ATP Imaging Using a Merocyanine-Hybrided Rhodamine Fluorescent Probe

The real-time intravital imaging of adenosine triphosphate (ATP) is essential for comprehending the intricate relationship between energy metabolism and cellular functions. To develop highly effective molecular tools for intravital ATP imaging, herein we designed a merocyanine-hybrided rhodamine fluorescent probe named PCR. The advantages offered by this probe toward ATP include near-infrared excitation/emission (λ_ex/λ_em = 700/758 nm), a rapid (<3 min) and reversible response, an ideal dissociation constant (K_d = 7.05 mM), a relatively low detection limit (36.9 μM), deep tissue penetration (180 μm), and two-photon excitability. With the aid of the probe, we successfully detected both exogenous and endogenous ATP generation in viable cells. Furthermore, we monitored dynamic fluctuations in ATP levels under various metabolic perturbations and observed an augmented ATP production in cancer cells relative to normal cells. Subsequently, our investigation unveiled that β-lapachone, a well-established anticancer agent, induces cancer cell death by triggering reactive oxygen species-dependent energy crisis. Lastly, we visualized oscillations in ATP levels during cerebral ischemia/reperfusion by examining HT-22 cells and live mouse brain. The aforementioned successful application showcases the probe's potential for further elucidating ATP function in intricate biological environments, particularly in vivo.