Calcium dynamics are important in the physiology of olfactory and vomeronasal sensory neurons (OSNs and VSNs). The focus of attention is primarily on Ca2+ influx, because Ca2+ carries the current of the cyclic nucleotide gated channel (CNGC) that opens upon odorant stimulation of the OSN and both internal stores and extracellular Ca 2+ contribute to the rise in intracellular Ca 2+ in VSNs upon stimulation with pheromone. Once Ca 2+ enters, the kinetics of its removal become important because, for example in OSNs, Ca 2+ can amplify the odor evoked currents by activating a CI conductance, amplify second messenger production, inhibit the CNGC, activate feed back regulation, inhibit key enzymes in signal transduction, among other effects. Therefore, removal of Ca2vand the kinetics of this removal can modulate the excitability and function of OSNs and VSNs, but less attention has been paid to Ca 2+ clearance than to increases. The Na/Ca exchanger plays a role in this Ca 2+ removal, but there is evidence from the past and our preliminary data that support a role for Plasma Membrane Calcium ATPases (Ca 2+ pumps, PMCAs) in this removal. We provide additional preliminary data that demonstrate the presence of PMCA isoforms in mouse OSNs and VNO. In particular, PMCAs 1 and 2 are found in OSNs and VNO, and PMCA2 may be localized to the OSN dendritic knob. These observations have implications for the functions of the PMCAs in OSNs and VSNs because each of the 4 PMCA isoforms has very different regulatory properties and affinities for Ca 2+. We propose three specific aims to partially test the hypotheses 1) PMCAs play a role in restoring calcium to basal levels after stimulation of OSNs and VSNs and 2) deletion of PMCA2 (in knock-out mice) leads to impairment in response to repetitive odor stimulation due to failure to return Ca 2v levels to basal in OSNs and VSNs.
Specific Aim 1. Identify the PMCA isoforms in the olfactory and VNO epithelia and their cellular Iocalizations on the OSNs and VSNS using Western blotting and immunohistochemistry of epithelia and immuno-fluorescence of dissociated cells.
Specific Aim 2. Test whether PMCAs among other Ca 2+ clearance mechanisms contribute to restoration of Ca2+ to basal levels after stimulation using calcium imaging of dissociated cells.
Specific Aim 3. Examine the OSNs and VSNs from PMCA2 knock-out mice using the techniques of Specific Aims 1 and 2. This exploratory work will improve our understanding of the contributions of PMCAs to OSN and VSN Ca 2+ clearance, give new insight into VSN and OSN physiology and transduction regulation, and open up new avenues for chemosensory research. The exploratory research will take advantage of the mouse genetic model and also allows the PI to apply her expertise in PMCAs from other chemosensory systems to the mouse. The understanding of OSN and VSN function is important for understanding normal olfactory and VNO function in health and disease.
| Weeraratne, S Dilhan; Valentine, Megan; Cusick, Matthew et al. (2006) Plasma membrane calcium pumps in mouse olfactory sensory neurons. Chem Senses 31:725-30 |
