RT Journal Article SR Electronic T1 Surface Traffic of Dendritic CaV1.2 Calcium Channels in Hippocampal Neurons JF The Journal of Neuroscience JO J. Neurosci. FD Society for Neuroscience SP 13682 OP 13694 DO 10.1523/JNEUROSCI.2300-11.2011 VO 31 IS 38 A1 Valentina Di Biase A1 Petronel Tuluc A1 Marta Campiglio A1 Gerald J. Obermair A1 Martin Heine A1 Bernhard E. Flucher YR 2011 UL http://www.jneurosci.org/content/31/38/13682.abstract AB In neurons L-type calcium currents function in gene regulation and synaptic plasticity, while excessive calcium influx leads to excitotoxicity and neurodegeneration. The major neuronal CaV1.2 L-type channels are localized in clusters in dendritic shafts and spines. Whereas CaV1.2 clusters remain stable during NMDA-induced synaptic depression, L-type calcium currents are rapidly downregulated during strong excitatory stimulation. Here we used fluorescence recovery after photobleaching (FRAP), live cell-labeling protocols, and single particle tracking (SPT) to analyze the turnover and surface traffic of CaV1.2 in dendrites of mature cultured mouse and rat hippocampal neurons, respectively. FRAP analysis of channels extracellularly tagged with superecliptic pHluorin (CaV1.2-SEP) demonstrated ∼20% recovery within 2 min without reappearance of clusters. Pulse–chase labeling showed that membrane-expressed CaV1.2-HA is not internalized within1 h, while blocking dynamin-dependent endocytosis resulted in increased cluster density after 30 min. Together, these results suggest a turnover rate of clustered CaV1.2s on the hour time scale. Direct recording of the lateral movement in the membrane using SPT demonstrated that dendritic CaV1.2s show highly confined mobility with diffusion coefficients of ∼0.005 μm2 s−1. Consistent with the mobile CaV1.2 fraction observed in FRAP, a ∼30% subpopulation of channels reversibly exchanged between confined and diffusive states. Remarkably, high potassium depolarization did not alter the recovery rates in FRAP or the diffusion coefficients in SPT analyses. Thus, an equilibrium of clustered and dynamic CaV1.2s maintains stable calcium channel complexes involved in activity-dependent cell signaling, whereas the minor mobile channel pool in mature neurons allows limited capacity for short-term adaptations.