Phoneutria nigriventer toxin Tx3-1 blocks A-type K+ currents controlling Ca2+ oscillation frequency in GH3 cells.
GH3 cells present spontaneous Ca2+ action potentials and oscillations of intracellular Ca2+, which can be modified by altering the activity of K+ or Ca2+ channels. We took advantage of this spontaneous activity to screen for effects of a purified toxin (Tx3-1) from the venom of Phoneutria nigriventer on ion channels. We report that Tx3-1 increases the frequency of Ca2+ oscillations, as do two blockers of potassium channels, 4-aminopyridine and charybdotoxin. Whole-cell patch clamp experiments show that Tx3-1 reversibly inhibits the A-type K+ current (I(A)) but does not block other K+ currents (delayed-rectifying, inward-rectifying, and large-conductance Ca2+-sensitive) or Ca2+ channels (T and L type) in these cells. In addition, we describe the sequence of a full cDNA clone of Tx3-1, which shows that Tx3-1 has no homology to other known blockers of K+ channels and gives insights into the processing of this neurotoxin. We conclude that Tx3-1 is a selective inhibitor of I(A), which can be used to probe the role of this channel in the control of cellular function. Based on the effect of Tx3-1, we suggest that I(A) is an important determinant of the frequency of Ca2+ oscillations in unstimulated GH3 cells.
Kushmerick
C
C
Departamento de Farmacologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Brasil.
Kalapothakis
E
E
Beirão
P S
PS
Penaforte
C L
CL
Prado
V F
VF
Cruz
J S
JS
Diniz
C R
CR
Cordeiro
M N
MN
Gomez
M V
MV
Romano-Silva
M A
MA
Prado
M A
MA
D015761:4-Aminopyridine / Q000494:pharmacology; D000595:Amino Acid Sequence; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000502:physiology; D002460:Cell Line; D018999:Charybdotoxin / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D018076:DNA, Complementary / Q000302:isolation & purification; D004533:Egtazic Acid / Q000494:pharmacology; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000235:genetics / Q000494:pharmacology*; D009543:Nifedipine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010507:Periodicity; D010902:Pituitary Gland / Q000737:chemistry / Q000166:cytology; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000502:physiology*; D013111:Spider Venoms / Q000235:genetics / Q000494:pharmacology*
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D002614:Chelating Agents; D018076:DNA, Complementary; D009479:Neuropeptides; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D013111:Spider Venoms; C098736:Tx3 neurotoxin; D018999:Charybdotoxin; D004533:Egtazic Acid; D015761:4-Aminopyridine; D009543:Nifedipine; D002118:Calcium