The rodent vomeronasal organ plays a crucial role in several social behaviors. Detection of pheromones or other emitted signaling molecules occurs in the dendritic microvilli of vomeronasal sensory neurons, where the binding of molecules to vomeronasal receptors leads to the influx of sodium and calcium ions mainly through the transient receptor potential canonical 2 (TRPC2) channel. To investigate the physiological role played by the increase in intracellular calcium concentration in the apical region of these neurons, we produced localized, rapid, and reproducible increases in calcium concentration with flash photolysis of caged calcium and measured calcium-activated currents with the whole cell voltage-clamp technique. On average, a large inward calcium-activated current of −261 pA was measured at −50 mV, rising with a time constant of 13 ms. Ion substitution experiments showed that this current is anion selective. Moreover, the chloride channel blockers niflumic acid and 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid partially inhibited the calcium-activated current. These results directly demonstrate that a large chloride current can be activated by calcium in the apical region of mouse vomeronasal sensory neurons. Furthermore, we showed by immunohistochemistry that the calcium-activated chloride channels TMEM16A/anoctamin1 and TMEM16B/anoctamin2 are present in the apical layer of the vomeronasal epithelium, where they largely colocalize with the TRPC2 transduction channel. Immunocytochemistry on isolated vomeronasal sensory neurons showed that TMEM16A and TMEM16B coexpress in the neuronal microvilli. Therefore, we conclude that microvilli of mouse vomeronasal sensory neurons have a high density of calcium-activated chloride channels that may play an important role in vomeronasal transduction.

Calcium calmodulin dependent kinase II (CaMKII) is sequestered in dendritic spines within seconds upon synaptic stimulation. The program Smoldyn was used to develop scenarios of single molecule CaMKII diffusion and binding in virtual dendritic spines. We first validated simulation of diffusion as a function of spine morphology. Additional cellular structures were then incorporated to simulate binding of CaMKII to the post-synaptic density (PSD); binding to cytoskeleton; or their self-aggregation. The distributions of GFP tagged native and mutant constructs in dissociated hippocampal neurons were measured to guide quantitative analysis. Intra-spine viscosity was estimated from fluorescence recovery after photo-bleach (FRAP) of red fluorescent protein. Intra-spine mobility of the GFP-CaMKIIα constructs was measured, with hundred-millisecond or better time resolution, from FRAP of distal spine tips in conjunction with fluorescence loss (FLIP) from proximal regions. Different FRAP \ FLIP profiles were predicted from our Scenarios and provided a means to differentiate binding to the PSDs from self-aggregation. The predictions were validated by experiments. Simulated fits of the Scenarios provided estimates of binding and rate constants. We utilized these values to assess the role of self-aggregation during the initial response of native CaMKII holoenzymes to stimulation. The computations revealed that self-aggregation could provide a concentration-dependent switch to amplify CaMKII sequestration and regulate its activity depending on its occupancy of the actin cytoskeleton.

TMEM16A is an essential component of Ca2+-activated Cl− currents in mouse vomeronasal sensory neurons.

The enzyme urease is an essential colonizing factor of the notorious carcinogenic pathogen Helicobacter pylori (H. pylori), conferring acid resistance to the bacterium.

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