ABSTRACT
Huntington's disease (HD) is a fatal, hereditary neurodegenerative disorder that causes chorea, cognitive deficits, and psychiatric symptoms. It is characterized by accumulation of mutant Htt protein, which primarily impacts striatal medium-sized spiny neurons (MSNs), as well as cortical pyramidal neurons (CPNs), causing synapse loss and eventually cell death. Perturbed Ca2+ homeostasis is believed to play a major role in HD, as altered Ca2+ homeostasis often precedes striatal dysfunction and manifestation of HD symptoms. In addition, dysregulation of Ca2+ can cause morphological and functional changes in MSNs and CPNs. Therefore, Ca2+ imaging techniques have the potential of visualizing changes in Ca2+ dynamics and neuronal activity in HD animal models. This minireview focuses on studies using diverse Ca2+ imaging techniques, including two-photon microscopy, fiber photometry, and miniscopes, in combination of Ca2+ indicators to monitor activity of neurons in HD models as the disease progresses. We then discuss the future applications of Ca2+ imaging to visualize disease mechanisms and alterations associated with HD, as well as studies showing how, as a proof-of-concept, Ca2+imaging using miniscopes in freely-behaving animals can help elucidate the differential role of direct and indirect pathway MSNs in HD symptoms.
ABSTRACT
Calcium-modulated photoactivatable ratiometric integrator (CaMPARI) is a calcium ion (Ca2+)- and light-dependent genetically encoded fluorescent activity integrator that can capture snapshots of neuronal activity through an irreversible process known as photoconversion. This unique property was previously used to label neurons based upon their tuning properties in order to map synaptic connectivity and to record large-scale neuronal activity in freely moving mice without attaching any mechanical device to them. The latest version of CaMPARI (CaMPARI2) was engineered to enhance the contrast generated by photoconverting the green protein to the activity-dependent red form and to reduce the Ca2+-independent photoconversion rate compared to the first generation of CaMPARI (CaMPARI1). However, here we show that this optimization process also resulted in reduced photoconversion efficiency of active neurons in the mouse cortex and hippocampus. Through side-by-side comparison of the two CaMPARI sensors under several experimental conditions, we show that CaMPARI1 exhibits a substantially higher red-to-green ratio in active cells than CaMPARI2. In addition, we show that CaMPARI1 also functions as a more sensitive traditional Ca2+ sensor than CaMPARI2 by producing larger activity-driven dynamic fluorescence changes in the observed neurons. Therefore, we conclude that during the optimization process of CaMPARI2, some of the sensor's characteristics were not predicted properly by in vitro screening assays, and therefore in vivo screening and validation steps should be included in future optimization attempts to increase the predictability of screening pipelines.