Protoporphyrins modulate voltage-gated Ca current in AtT-20 pituitary cells

Abstract

1. Zinc-protoporphyrin-IX (ZnPP-IX) is an inhibitor of the enzyme heme-oxygenase-2 (HO-2) and consequently has been used to examine the role of carbon monoxide production in neural tissues. We have measured voltage-gated Ca current in AtT-20 pituitary cells using the whole-cell patch-clamp technique and have assessed the effects of extracellularly applied ZnPP-IX and related compounds. 2. Ca currents evoked by depolarizing steps from a holding potential of -90 mV were of the high-threshold, slowly inactivating type. Fifty-six percent of this current was blocked by 10 microM nifedipine and 16% by 3 microM omega-conotoxin with the remainder resistant to both drugs in combination, suggesting that the total Ca current was a mixture of L, N, and possibly P-type conductances. 3. Bath application of ZnPP-IX resulted in an irreversible, dose-dependent attenuation of Ca current. Five micromolar ZnPP-IX produced a 62% reduction of peak current amplitude with no shift in the current-voltage relation, 0.5 microM produced a 19% reduction, and 0.05 microM produced a variable response, either a small transient attenuation or potentiation. 4. The attenuation of Ca current by 5 microM ZnPP-IX could be nearly completely blocked by co-application of superoxide dismutase in the bath (90 U/ml) but not by addition of an inhibitor of cGMP-dependent protein kinase to the internal saline (KT5823, 1 microM). 5. Other inhibitors of heme-oxygenase with similar potency such as tin-protoporphyrin-IX (Sn-PP-IX) and Zn-deuteroporphyrin-bis-glycol (ZnBG) did not attenuate Ca current when applied at 5microM.(ABSTRACT TRUNCATED AT 400 WORDS)

Carbon Monoxide: Endogenous Production, Physiological Functions, and Pharmacological Applications

“…Extracellular application of ZnPP in AtT-20 pituitary cells irreversibly inhibited the total Ca 2ϩ currents conducted by L-, N-, and possibly P-type voltage-gated Ca 2ϩ channels (Linden et al, 1993). However, the effect of ZnPP largely resulted from the generation of superoxide anion, instead of HO inhibition.…”

Carbon monoxide donors or heme oxygenase-1 (HO-1) overexpression blocks interleukin-18-mediated NF-κB–PTEN-dependent human cardiac endothelial cell death

“…While CoPPIX acts a CO donor, ZnPPIX serves as an inhibitor. ZnPPIX has also been shown to irreversibly attenuate Ca 2+ currents [22], inhibit hematopoisis in rabbit and human bone marrow [23], and non-selectively inhibit NOS enzymes and soluble guanylate cyclase [24]. Both CoPPIX and ZnPPIX were dissolved in 0.2M NaOH, neutralized with 1M HCl, adjusted to 1 mg/ml concentration, filter sterilized and stored at −80°C until further use.…”

Unique Properties of Polyphenol Stilbenes in the Brain: More than Direct Antioxidant Actions; Gene/Protein Regulatory Activity

“…In that it is a gas, carbon monoxide can travel freely through intracellular and extracellular compartments. The CO literature is vast and complex with many controversies that have yet to be resolved [58][59][60][61][62][63] . CO is better known to be toxic at high levels.…”