Molecular alterations in the cerebellum of the plasma membrane calcium ATPase 2 (PMCA2)-null mouse indicate abnormalities in Purkinje neurons

Abstract

PMCA2, a major calcium pump, is expressed at particularly high levels in Purkinje neurons. Accordingly, PMCA2-null mice exhibit ataxia suggesting cerebellar pathology. It is not yet known how changes in PMCA2 expression or activity affect molecular pathways in Purkinje neurons. We now report that the levels of metabotropic glutamate receptor 1 (mGluR1), which plays essential roles in motor coordination, synaptic plasticity, and associative learning, are reduced in the cerebellum of PMCA2-null mice as compared to wild type littermates. The levels of inositol 1,4,5-triphosphate receptor type 1 (IP3R1), an effector downstream to mGluR1, which mediates intracellular calcium signaling, and the expression of Homer 1b/c and Homer 3, scaffold proteins that couple mGluR1 to IP3R1, are also reduced in somata and dendrites of some Purkinje cell subpopulations. In contrast, no alterations occur in the levels of mGluR1 and its downstream effectors in the hippocampus, indicating that the effects are region specific. The reduction in cerebellar mGluR1, IP3R1 and Homer 3 levels are neither due to a generic decrease in Purkinje proteins nor extensive dendritic loss as immunoreactivity to total and non-phosphorylated neurofilament H (NFH) is increased in Purkinje dendrites and microtubule associated protein 2 (MAP2) staining reveals a dense dendritic network in the molecular layer of the PMCA2-null mouse cerebellum. PMCA2 coimmunoprecipitates with mGluR1, Homer 3 and IP3R1, suggesting that the calcium pump is a constituent of the mGluR1 signaling complex. Our results suggest that the decrease in the expression of mGluR1 and its downstream effectors and perturbations in the mGluR1 signaling complex in the absence of PMCA2 may cumulatively result in aberrant metabotropic glutamate receptor signaling in Purkinje neurons leading to cerebellar deficits in the PMCA2-null mouse.

Plasma Membrane Ca2+ATPase 2 Contributes to Short-Term Synapse Plasticity at the Parallel Fiber to Purkinje Neuron Synapse

“…The high levels of PMCA2 in the PN also suggest an important contribution from Ca 2ϩ extrusion during the developmental shaping of the PN dendritic tree, especially at a time of intense synaptogenesis, when PMCA2 levels rise (Jensen et al, 2004). Alternatively, recent reports that the PMCA2Ϫ/Ϫ cerebellum expresses fewer homer/metabotropic glutamate receptor 1 (mGluR1)/IP 3 -receptor (IP 3 R) complexes (Kurnellas et al, 2006) and that loss of IP 3 -R-mediated signaling through mGluR1 activation in GCs prevents BDNF-mediated outgrowth of PNs (Hisatsune et al, 2006) provides a possible explanation for the stunted appearance of the PN dendritic tree in PMCA2Ϫ/Ϫ mice.…”

Cytosolic Ca2+ Buffers

“…This is indicative of a Ca v channel down-regulation likely to regulate [Ca 2þ ] i toward normal homeostasis (Ueno et al 2002). PMCA2 2/2 mice have decreased expression levels of CB-D28k (Hu et al 2006), metabotropic glutamate receptor 1 (mGluR1), and of InsP 3 receptor type 1 (IP3R1), responsible for the Ca 2þ release from ER stores (Kurnellas et al 2007) (Fig. 2).…”

Contribution of plasma membrane Ca2+ATPase to cerebellar synapse function

“…This is a very interesting finding and provided the first evidence of a signalling role for a PMCA2 molecular interaction in a post-synaptic cerebellar compartment, additional to its role in calcium clearance [25] . Subsequent electrophysiological analysis shows that the mGluR1 current is markedly reduced in the PMCA2 -/-cerebellar PNs (Figure 4), even though the mGluR1 receptors could be detected, if disorganized, in these cells [24,27] . The reduction in mGluR1 mediated synaptic signalling could provide one of the most important clues as to the behavioural cerebellar ataxia in the PMCA2 -/-mice.…”

“…A recent report also indicated that PMCA2 contributes to the post-synaptic molecular complex formed between the InsP3 receptor, metabotropic glutamate receptors (mGluR1) and homer, a signalling adaptor scaffolding protein [24] . This is a very interesting finding and provided the first evidence of a signalling role for a PMCA2 molecular interaction in a post-synaptic cerebellar compartment, additional to its role in calcium clearance [25] .…”

“…-/-mice. Given the interaction between metabotropic glutamate receptors (mGluR1) and PMCA2 [24] we tested mGluR1 function using an electrophysiological assay. A: Representative traces of mGluR1 dependent inward currents in response to increasing numbers of 100 Hz stimulation to the PFs [36] .…”