Modulation of CaV3.2 T-type calcium channels by GM1

KEELING G; SHANKARAPPA S. A; WEISSMANN C; PERISSINOTTI PP; CRIBBS L; STUBBS EB JR; PIEDRAS-RENTERÍA ES

Washington, DC

Congreso; Society for Neuroscience; 2014

Gangliosides are sialic acid-containing, complex glycosphingolipids naturally enriched in neuronal membranes. They are important in the regulation of signaling protein function, cell morphology, cell cycle and neurotransmission; and they also function as membrane receptors for compounds such as cholera, tetanus and botulinum toxin. Alterations in ganglioside concentration are associated with gangliosiodoses. The monosyaloganglioside GM1 is particularly abundant in the plasma membrane of neurons. The segregation of gangliosides within restricted lipid rafts provides the molecular basis for lateral interactions between them and membrane proteins. GM1 is a known modulator of L-type calcium channels in neuroblastoma cells (Carlson et al, 1994) and cerebellar granule neurons (Wu et al, 1996); and of CaV2.3 channels in sperm (Cohen et al, 2014). Here we report the modulation of CaV3.2 channels by GM1. We analyzed the effect of GM1 on CaV3.2 channels constitutively expressed in HEK293 cells. Treatment with 1 µM GM1 decreased current densities by 32%, an effect that could be completely reversed by addition of the B subunit of Cholera toxin (ChTxB), a specific binding ligand for GM1. This effect was specific for CaV3.2, as GM1 had no effect on currents arising from CaV3.1. Moreover, GD1a, a ganglioside with similar structure to GM1 induced no changes in CaV3.2 current densities. Treatment with GM1 also resulted in a hyperpolarizing shift in steady state inactivation, not reverted by ChTxB. Co-localization analysis exhibits a strong association between CaV3.2 and GM1 (Pearson?s coefficient 84 ± 0.02 %), yielding a Manders? coefficient of 75 ± 0.03 % of the GM1 signal associated with CaV3.2. Hyperglycemic treatment (25 mM glucose) in culture for 24 hrs resulted in a reduction of GM1-induced inhibition due to a decreased ganglioside presence. Our studies show that GM1 is an endogenous, reversible, negative modulator of T-type CaV3.2 channels and that conditions leading to changes in GM1 levels at the membrane may affect neuronal excitability and could contribute to the beneficial effects of GM1 therapies. Dis-inhibition of CaV3.2 channels by GM1 could contribute to increased excitability in diabetic neuropathy and other diseases.