The K+ channel openers diazoxide and NS1619 induce depolarization of mitochondria and have differential effects on cell Ca2+ in CD34+ cell line KG-1a.
OBJECTIVE
Mitochondrial membrane potential (deltaPsim) and intracellular Ca2+ play a crucial role in growth and differentiation in hemopoiesis. Some potassium channel openers such as diazoxide have the capacity to elevate cytosolic Ca2+ and depolarize mitochondria in cardiomyocytes. To clarify if such substances have effects on hemopoietic cells we investigated the commonly used opener of the mitoK(ATP) channel, diazoxide, and the opener of BK channels, NS1619, for their potential to depolarize mitochondria, elevate cytosolic Ca2+, and induce apoptosis in the hemopoietic CD34+ cell line KG-1a.


METHODS
Fluorescent probes were used to investigate deltaPsim, free Ca2+, and apoptosis (JC-1, fluo-3-AM and annexin V-FITC) by flow cytometry. To measure deltaPsim with JC-1 in glycoprotein P+ cells we used an improved dye loading technique with verapamil.


RESULTS
NS1619 induced stronger dose-dependent mitochondrial depolarizations than diazoxide. Depolarization was independent from caspase activation and could also be induced when the driving force for K+ out of cells was near 0 mV. In Ca2+ free solutions NS1619 induced stronger Ca2+ elevations than diazoxide and elevated Ca2+ also after Ca2+ depletion of the endoplasmatic reticulum with caffeine. NS1619 did not enhance the Ca2+ elevation induced by ionophores (CCCP, valinomycin) that depolarize mitochondria. Both agents were weak inducers of apoptosis.


CONCLUSIONS
Diazoxide has similar effects in CD34+ cells as described for muscle or nerve cells. In accordance to the single channel conductance of mitoK(ATP) and BK channels, NS1619 is a more potent inducer of mitochondrial depolarization than diazoxide. NS1619 releases Ca2+ from an intracellular pool that is insensitive to caffeine but depends strongly on deltaPsim.
Körper
Sixten
S
Freie Universität Berlin, Universitätsklinikum Benjamin Franklin, Medizinische Klink III (Hämatologie, Onkologie und Transfusionsmedizin), Berlin, Germany. sixten.koerper@medizin.uni-ulm.de
Nolte
Florian
F
Rojewski
Markus Thomas
MT
Thiel
Eckhard
E
Schrezenmeier
Hubert
H
D018952:Antigens, CD34; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D001562:Benzimidazoles / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D003981:Diazoxide / Q000494:pharmacology*; D005456:Fluorescent Dyes; D018922:HL-60 Cells; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects*; D008928:Mitochondria / Q000187:drug effects* / Q000502:physiology*; D015221:Potassium Channels / Q000819:agonists / Q000502:physiology*; D014665:Vasodilator Agents / Q000494:pharmacology*
D018952:Antigens, CD34; D001562:Benzimidazoles; D005456:Fluorescent Dyes; D015221:Potassium Channels; D014665:Vasodilator Agents; C086491:NS 1619; D003981:Diazoxide; D002118:Calcium