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Inhibitory modulation of fast and slow Ca(2+)-currents in neuroblastoma x glioma cells during differentiation
1II. Physiologisches Institut der Universität des Saarlandes, Homburg, F.R.G.
Abstract:
Mouse neuroblastoma x rat glioma hybrid cells (N x G, 108CC15) were used to study the inhibitory effects of the synthetic opioid D-Ala2-D-Leu5-enkephalin (DADLE), somatostatin, adrenaline-alpha 2 and angiotensin II on voltage-dependent Ca(2+)-currents (ICa) using the patch-clamp technique in the whole-cell configuration mode. The inhibitory effects could be abolished by pretreatment of N x G cells with pertussis toxin or intracellular infusion of GDP beta S indicating an involvement of a pertussis toxin sensitive GTP-binding protein (G-protein), presumably Go. The effect of DADLE, the strongest inhibitor of ICa, was studied during dibutyryl cyclic AMP (dBcAMP) induced differentiation. Using omega-conotoxin GVIA (omega-CTX) and methoxyverapamil (D600) as specific Ca(2+)-channel blockers of the N- and L-type Ca(2+)-channels, it was found that in N x G cells DADLE predominantly induces inhibition of T- and N-type Ca(2+)-channels.
Insights
Synthetic opioids like DADLE inhibit voltage-dependent calcium currents (ICa) in hybrid N x G cells. This inhibition involves a specific G-protein and primarily affects T- and N-type calcium channels.
Area of Science:
- Neuroscience
- Cell Biology
- Pharmacology
Background:
- Voltage-dependent calcium currents (ICa) play crucial roles in neuronal function.
- Hybrid neuroblastoma x glioma cells (N x G) provide a model for studying neuronal signaling pathways.
- G-protein coupled receptors mediate diverse cellular responses, including modulation of ion channels.
Purpose of the Study:
- To investigate the inhibitory effects of D-Ala2-D-Leu5-enkephalin (DADLE), somatostatin, adrenaline-alpha 2, and angiotensin II on voltage-dependent Ca(2+)-currents (ICa) in N x G cells.
- To elucidate the involvement of G-proteins in mediating these inhibitory effects.
- To determine the specific types of Ca(2+) channels affected by DADLE.
Main Methods:
- Patch-clamp technique in whole-cell configuration was employed to measure ICa.
- Cells were pretreated with pertussis toxin or infused with GDP beta S to assess G-protein involvement.
- Specific calcium channel blockers, omega-conotoxin GVIA (omega-CTX) and methoxyverapamil (D600), were used to identify affected channel subtypes.
- Dibutyryl cyclic AMP (dBcAMP) was used to induce cellular differentiation.
Main Results:
- DADLE, somatostatin, adrenaline-alpha 2, and angiotensin II inhibited ICa in N x G cells.
- Inhibitory effects were abolished by pertussis toxin or GDP beta S, indicating involvement of a pertussis toxin-sensitive G-protein (likely Go).
- DADLE predominantly inhibited T- and N-type Ca(2+) channels, as evidenced by experiments with omega-CTX and D600.
Conclusions:
- A pertussis toxin-sensitive G-protein mediates the inhibitory effects of DADLE, somatostatin, adrenaline-alpha 2, and angiotensin II on ICa in N x G cells.
- DADLE exerts its inhibitory action primarily on T- and N-type calcium channels.
- These findings contribute to understanding the regulation of calcium channels by G-protein coupled receptors in neuronal cells.