L-type calcium channel blockers and substance P induce angiogenesis of cortical vessels associated with beta-amyloid plaques in an Alzheimer mouse model

AbstractIt is well established that L-type calcium channels (LTCCs) are expressed in astroglia. However, their functional role is still speculative, especially under pathologic conditions. We recently showed that the α1 subunit-like immunoreactivity of the CaV1.2 channel is strongly expressed in reactive astrocytes around beta-amyloid plaques in 11-month-old Alzheimer transgenic (tg) mice with the amyloid precursor protein London and Swedish mutations. The aim of the present study was to examine the cellular expression of all LTCC subunits around beta-amyloid plaques by in situ hybridization using 35S-labeled oligonucleotides. Our data show that messenger RNAs (mRNAs) of the LTCC CaV1.2 α1 subunit as well as all auxiliary β and α2δ subunits, except α2δ-4, were expressed in the hippocampus of age-matched wild-type mice. It was unexpected to see, that cells directly located in the plaque core in the cortex expressed mRNAs for CaV1.2 α1, β2, β4, and α2δ-1, whereas no expression was detected in the halo. Furthermore, cells in the plaque core also expressed preprotachykinin-A mRNA, the precursor for substance P. By means of confocal microscopy, we demonstrated that collagen-IV-stained brain vessels in the cortex were associated with the plaque core and were immunoreactive for substance P. In cortical organotypic brain slices of adult Alzheimer mice, we could demonstrate that LTCC blockers increased angiogenesis, which was further potentiated by substance P. In conclusion, our data show that brain vessels associated with beta-amyloid plaques express substance P and an LTCC and may play a role in angiogenesis

Densin-180 controls the trafficking and signaling of L-type voltage-gated Ca_v 1.2 Ca^(2+) channels at excitatory synapses

Voltage-gated Ca_v1.2 and Ca_v1.3 (L-type) Ca^(2+) channels regulate neuronal excitability, synaptic plasticity, and learning and memory. Densin-180 (densin) is an excitatory synaptic protein that promotes Ca^(2+)-dependent facilitation of voltage-gated Ca_v1.3 Ca^(2+) channels in transfected cells. Mice lacking densin (densin KO) exhibit defects in synaptic plasticity, spatial memory, and increased anxiety-related behaviors --phenotypes that more closely match those in mice lacking Ca_v1.2 than Ca_v1.3. Thus, we investigated the functional impact of densin on Ca_v1.2. We report that densin is an essential regulator of Ca_v1.2 in neurons, but has distinct modulatory effects compared to its regulation of Ca_v1.3. Densin binds to the N-terminal domain of Ca_v1.2 but not Ca_v1.3, and increases Ca_v1.2 currents in transfected cells and in neurons. In transfected cells, densin accelerates the forward trafficking of Ca_v1.2 channels without affecting their endocytosis. Consistent with a role for densin in increasing the number of postsynaptic Ca_v1.2 channels, overexpression of densin increases the clustering of Ca_v1.2 in dendrites of hippocampal neurons in culture. Compared to wild-type mice, the cell-surface levels of Ca_v1.2 in the brain as well as Ca_v1.2 current density and signaling to the nucleus are reduced in neurons from densin KO mice. We conclude that densin is an essential regulator of neuronal Ca_v1 channels and ensures efficient Ca_v1.2 Ca^(2+) signaling at excitatory synapses

Restricting calcium currents is required for correct fiber type specification in skeletal muscle

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α2δ-4 and Cachd1 proteins are regulators of presynaptic functions

The α2δ auxiliary subunits of voltage-gated calcium channels (VGCC) were traditionally regarded as modulators of biophysical channel properties. In recent years, channel-independent functions of these subunits, such as involvement in synapse formation, have been identified. In the central nervous system, α2δ isoforms 1, 2, and 3 are strongly expressed, regulating glutamatergic synapse formation by a presynaptic mechanism. Although the α2δ-4 isoform is predominantly found in the retina with very little expression in the brain, it was recently linked to brain functions. In contrast, Cachd1, a novel α2δ-like protein, shows strong expression in brain, but its function in neurons is not yet known. Therefore, we aimed to investigate the presynaptic functions of α2δ-4 and Cachd1 by expressing individual proteins in cultured hippocampal neurons. Both α2δ-4 and Cachd1 are expressed in the presynaptic membrane and could rescue a severe synaptic defect present in triple knockout/knockdown neurons that lacked the α2δ-1-3 isoforms (α2δ TKO/KD). This observation suggests that presynaptic localization and the regulation of synapse formation in glutamatergic neurons is a general feature of α2δ proteins. In contrast to this redundant presynaptic function, α2δ-4 and Cachd1 differentially regulate the abundance of presynaptic calcium channels and the amplitude of presynaptic calcium transients. These functional differences may be caused by subtle isoform-specific differences in α1 -α2 δ protein–protein interactions, as revealed by structural homology modelling. Taken together, our study identifies both α2δ-4 and Cachd1 as presynaptic regulators of synapse formation, differentiation, and calcium channel functions that can at least partially compensate for the loss of α2δ-1-3. Moreover, we show that regulating glutamatergic synapse formation and differentiation is a critical and surprisingly redundant function of α2δ and Cachd1

Presynaptic α2δ subunits are key organizers of glutamatergic synapses

In nerve cells the genes encoding for α2δ subunits of voltage-gated calcium channels have been linked to synaptic functions and neurological disease. Here we show that α2δ subunits are essential for the formation and organization of glutamatergic synapses. Using a cellular α2δ subunit triple-knockout/knockdown model, we demonstrate a failure in presynaptic differentiation evidenced by defective presynaptic calcium channel clustering and calcium influx, smaller presynaptic active zones, and a strongly reduced accumulation of presynaptic vesicle-associated proteins (synapsin and vGLUT). The presynaptic defect is associated with the downscaling of postsynaptic AMPA receptors and the postsynaptic density. The role of α2δ isoforms as synaptic organizers is highly redundant, as each individual α2δ isoform can rescue presynaptic calcium channel trafficking and expression of synaptic proteins. Moreover, α2δ-2 and α2δ-3 with mutated metal ion-dependent adhesion sites can fully rescue presynaptic synapsin expression but only partially calcium channel trafficking, suggesting that the regulatory role of α2δ subunits is independent from its role as a calcium channel subunit. Our findings influence the current view on excitatory synapse formation. First, our study suggests that postsynaptic differentiation is secondary to presynaptic differentiation. Second, the dependence of presynaptic differentiation on α2δ implicates α2δ subunits as potential nucleation points for the organization of synapses. Finally, our results suggest that α2δ subunits act as transsynaptic organizers of glutamatergic synapses, thereby aligning the synaptic active zone with the postsynaptic density

Cardiac-type EC-Coupling in Dysgenic Myotubes Restored with Ca(2+) Channel Subunit Isoforms α(1C) and α(1D) Does not Correlate with Current Density

Ca(2+)-induced Ca(2+)-release (CICR)—the mechanism of cardiac excitation-contraction (EC) coupling—also contributes to skeletal muscle contraction; however, its properties are still poorly understood. CICR in skeletal muscle can be induced independently of direct, calcium-independent activation of sarcoplasmic reticulum Ca(2+) release, by reconstituting dysgenic myotubes with the cardiac Ca(2+) channel α(1C) (Ca(V)1.2) subunit. Ca(2+) influx through α(1C) provides the trigger for opening the sarcoplasmic reticulum Ca(2+) release channels. Here we show that also the Ca(2+) channel α(1D) isoform (Ca(V)1.3) can restore cardiac-type EC-coupling. GFP-α(1D) expressed in dysgenic myotubes is correctly targeted into the triad junctions and generates action potential-induced Ca(2+) transients with the same efficiency as GFP-α(1C) despite threefold smaller Ca(2+) currents. In contrast, GFP-α(1A), which generates large currents but is not targeted into triads, rarely restores action potential-induced Ca(2+) transients. Thus, cardiac-type EC-coupling in skeletal myotubes depends primarily on the correct targeting of the voltage-gated Ca(2+) channels and less on their current size. Combined patch-clamp/fluo-4 Ca(2+) recordings revealed that the induction of Ca(2+) transients and their maximal amplitudes are independent of the different current densities of GFP-α(1C) and GFP-α(1D). These properties of cardiac-type EC-coupling in dysgenic myotubes are consistent with a CICR mechanism under the control of local Ca(2+) gradients in the triad junctions