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The C. elegans T-type calcium channel CCA-1 boosts neuromuscular transmission.

ABSTRACT: Low threshold-activated or T-type calcium channels are postulated to mediate a variety of bursting and rhythmic electrical firing events. However, T-type channels' exact physiological contributions have been difficult to assess because of their incompletely defined pharmacology and the difficulty in isolating T-type currents from more robust high threshold calcium currents. A current in C. elegans pharyngeal muscle displays the kinetic features of a T-type calcium channel and is absent in animals homozygous for mutations at the cca-1 locus (see accompanying paper). cca-1 is expressed in pharyngeal muscle and encodes a protein (CCA-1) with strong homology to the alpha1 subunits of vertebrate T-type channels. We show that CCA-1 plays a critical role at the pharyngeal neuromuscular junction, permitting the efficient initiation of action potentials in response to stimulation by the MC motor neuron. Loss of cca-1 function decreases the chance that excitatory input from MC will successfully trigger an action potential, and reduces the ability of an animal to take in food. Intracellular voltage recordings demonstrate that when wild-type cca-1 is absent, the depolarizing phase of the pharyngeal action potential tends to plateau or stall near -30 mV, the voltage at which the CCA-1 channel is likely to be activated. We conclude that the CCA-1 T-type calcium channel boosts the excitatory effect of synaptic input, allowing for reliable and rapid depolarization and contraction of the pharyngeal muscle. We also show that the pharyngeal muscle employs alternative strategies for initiating action potentials in certain cases of compromised MC motor neuron function.

SUBMITTER: Steger KA

PROVIDER: S-EPMC1382270 | biostudies-literature | 2005 Jun

REPOSITORIES: biostudies-literature

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The C. elegans T-type calcium channel CCA-1 boosts neuromuscular transmission.

Steger Katherine A KA Shtonda Boris B BB Thacker Colin C Snutch Terrance P TP Avery Leon L

The Journal of experimental biology 20050601 Pt 11

Low threshold-activated or T-type calcium channels are postulated to mediate a variety of bursting and rhythmic electrical firing events. However, T-type channels' exact physiological contributions have been difficult to assess because of their incompletely defined pharmacology and the difficulty in isolating T-type currents from more robust high threshold calcium currents. A current in C. elegans pharyngeal muscle displays the kinetic features of a T-type calcium channel and is absent in animal ...[more]

PMID: 15914662

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Project description:BackgroundThe conserved Caenorhabditis elegans proteins NID-1/nidogen and PTP-3A/LAR-RPTP function to efficiently localize the presynaptic scaffold protein SYD-2/?-liprin at active zones. Loss of function in these molecules results in defects in the size, morphology and spacing of neuromuscular junctions.ResultsHere we show that the Cav2-like voltage-gated calcium channel (VGCC) proteins, UNC-2 and UNC-36, and the calmodulin kinase II (CaMKII), UNC-43, function to regulate the size and morphology of presynaptic domains in C. elegans. Loss of function in unc-2, unc-36 or unc-43 resulted in slightly larger GABAergic neuromuscular junctions (NMJs), but could suppress the synaptic morphology defects found in nid-1/nidogen or ptp-3/LAR mutants. A gain-of-function mutation in unc-43 caused defects similar to those found in nid-1 mutants. Mutations in egl-19, Cav1-like, or cca-1, Cav3-like, ?1 subunits, or the second ?2/? subunit, tag-180, did not suppress nid-1, suggesting a specific interaction between unc-2 and the synaptic extracellular matrix (ECM) component nidogen. Using a synaptic vesicle marker in time-lapse microscopy studies, we observed GABAergic motor neurons adding NMJ-like structures during late larval development. The synaptic bouton addition appeared to form in at least two ways: (1) de novo formation, where a cluster of vesicles appeared to coalesce, or (2) when a single punctum became enlarged and then divided to form two discrete fluorescent puncta. In comparison to wild type animals, we found unc-2 mutants exhibited reduced NMJ dynamics, with fewer observed divisions during a similar stage of development.ConclusionsWe identified UNC-2/UNC-36 VGCCs and UNC-43/CaMKII as regulators of C. elegans synaptogenesis. UNC-2 has a modest role in synapse formation, but a broader role in regulating dynamic changes in the size and morphology of synapses that occur during organismal development. During the late 4th larval stage (L4), wild type animals exhibit synaptic morphologies that are similar to those found in animals lacking NID-1/PTP-3 adhesion, as well as those with constitutive activation of UNC-43. Genetic evidence indicates that the VGCCs and the NID-1/PTP-3 adhesion complex provide opposing functions in synaptic development, suggesting that modulation of synaptic adhesion may underlie synapse development in C. elegans.

Project description:Calcium homeostasis is critical for cardiac myocyte function and must be tightly regulated. The guiding hypothesis of this study is that a carboxyl-terminal cleavage product of the cardiac L-type calcium channel (Ca(V)1.2) autoregulates expression. First, we confirmed that the Ca(V)1.2 C terminus (CCt) is cleaved in murine cardiac myocytes from mature and developing ventricle. Overexpression of full-length CCt caused a 34+/-8% decrease of Ca(V)1.2 promoter activity, and truncated CCt caused an 80+/-3% decrease of Ca(V)1.2 promoter (n=12). The full-length CCt distributes into cytosol and nucleus. A deletion mutant of CCt has a greater relative affinity for the nucleus than full-length CCt, and this is consistent with increased repression of Ca(V)1.2 promoter activity by truncated CCt. Chromatin immunoprecipitation analysis revealed that CCt interacts with the Ca(V)1.2 promoter in adult ventricular cardiac myocytes at promoter modules containing Nkx2.5/Mef2, C/EBp, and a cis regulatory module. The next hypothesis tested was that CCt contributes to transcriptional signaling associated with cellular hypertrophy. We explored whether fetal cardiac myocyte Ca(V)1.2 was regulated by serum in vitro. We tested atrial natriuretic factor promoter activity as a positive control and measured the serum response of Ca(V)1.2 promoter, protein, and L-type current (I(Ca,L)) from fetal mouse ventricular myocytes. Serum increased atrial natriuretic factor promoter activity and cell size as expected. Serum withdrawal increased Ca(V)1.2 promoter activity, mRNA, and I(Ca,L). Moreover, serum withdrawal decreased the relative nuclear localization of CCt. A combination of promoter deletion mutant analyses, and the response of promoter mutants to serum withdrawal support the conclusion that CCt, a proteolytic fragment of Ca(V)1.2, autoregulates Ca(V)1.2 expression in cardiac myocytes. These data support the novel mechanism that a mobile segment of Ca(V)1.2 links Ca handling to nuclear signaling.

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The C. elegans T-type calcium channel CCA-1 boosts neuromuscular transmission.

Publications

The C. elegans T-type calcium channel CCA-1 boosts neuromuscular transmission.

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