Dynamic Ca²⁺ signals reflect acute changes in membrane excitability, and also mediate signaling cascades in chronic processes. In both cases, chronic Ca²⁺ imaging is often desired, but challenged by the cytotoxicity intrinsic to calmodulin (CaM)-based GCaMP, a series of genetically-encoded Ca²⁺ indicators that have been widely applied. Here, we demonstrate the performance of GCaMP-X in chronic Ca²⁺ imaging of cortical neurons, where GCaMP-X by design is to eliminate the unwanted interactions between the conventional GCaMP and endogenous (apo)CaM-binding proteins. By expressing in adult mice at high levels over an extended time frame, GCaMP-X showed less damage and improved performance in two-photon imaging of sensory (whisker-deflection) responses or spontaneous Ca²⁺ fluctuations, in comparison with GCaMP. Chronic Ca²⁺ imaging of one month or longer was conducted for cultured cortical neurons expressing GCaMP-X, unveiling that spontaneous/local Ca²⁺ transients progressively developed into autonomous/global Ca²⁺ oscillations. Along with the morphological indices of neurite length and soma size, the major metrics of oscillatory Ca²⁺, including rate, amplitude and synchrony were also examined. Dysregulations of both neuritogenesis and Ca²⁺ oscillations became discernible around 2–3 weeks after virus injection or drug induction to express GCaMP in newborn or mature neurons, which were exacerbated by stronger or prolonged expression of GCaMP. In contrast, neurons expressing GCaMP-X were significantly less damaged or perturbed, altogether highlighting the unique importance of oscillatory Ca²⁺ to neural development and neuronal health. In summary, GCaMP-X provides a viable solution for Ca²⁺ imaging applications involving long-time and/or high-level expression of Ca²⁺ probes.

The automatic initiation of actions can be highly functional. But occasionally these actions cannot be withheld and are released at inappropriate times, impulsively. Striatal activity has been shown to participate in the timing of action sequence initiation and it has been linked to impulsivity. Using a self-initiated task, we trained adult male rats to withhold a rewarded action sequence until a waiting time interval has elapsed. By analyzing neuronal activity we show that the striatal response preceding the initiation of the learned sequence is strongly modulated by the time subjects wait before eliciting the sequence. Interestingly, the modulation is steeper in adolescent rats, which show a strong prevalence of impulsive responses compared to adults. We hypothesize this anticipatory striatal activity reflects the animals’ subjective reward expectation, based on the elapsed waiting time, while the steeper waiting modulation in adolescence reflects age-related differences in temporal discounting, internal urgency states, or explore–exploit balance.