Activity labeling in vivo using CaMPARI2 reveals electrophysiological differences between neurons with high and low firing rate set points

Individual excitatory neurons in visual cortex (V1) display remarkably stable mean firing rates over many days, even though these rates can differ by several orders of magnitude between neurons. When perturbed, each neuron’s firing rate is slowly regulated back to its pre-perturbation level, demonstrating that neurons maintain their mean firing rate around an individual firing rate set point (FRSP). To better understand the mechanisms that neurons within a single cell type use to maintain different FRSPs in vivo, we implemented a novel method of activity labeling that uses CaMPARI2, a fluorescent protein that undergoes Ca2+- and UV-dependent green-to-red photoconversion, to permanently label neurons in freely behaving mice based on their firing rates. We found that immediate early gene (IEG) expression was correlated with CaMPARI2 red/green ratio following an activity stimulation paradigm, and that neurons with greater photoconversion in vivo tended to have a higher firing rate ex vivo. In layer 4 (L4) pyramidal neurons in mouse monocular V1, which comprise a single transcriptional cell type, we found that high activity neurons had a left-shifted F-I curve, lower rheobase current, and decreased spike adaptation index relative to low activity neurons, demonstrating increased intrinsic excitability. Surprisingly, we found no difference in total excitatory or inhibitory synaptic current or in E/I ratio between high and low activity neurons. Thus, within a single cell type differences in intrinsic excitability and spike frequency adaptation can contribute to divergent activity set points. These results reveal that E/I ratio plays only a minor role in determining the firing rate set point of L4 pyramidal neurons, while intrinsic excitability is an important factor.