Rat melanotrophs express several types of voltage-gated and ligand-gated calcium channels, although mechanisms involved in the maintenance of the resting intracellular Ca2+ concentration ([Ca2+]i) remain unknown. We analyzed mechanisms regulating resting [Ca2+]i in dissociated rat melanotrophs by Ca2+-imaging and patch-clamp techniques. Treatment with antagonists of L-type, but not N- or P/Q-type voltage-gated Ca2+ channels (VGCCs) as well as removal of extracellular Ca2+ resulted in a rapid and reversible decrease in [Ca2+]i, indicating constitutive Ca2+ influx through L-type VGCCs. Reduction of extracellular Na+ concentration (replacement with NMDG+) similarly decreased resting [Ca2+]i. When cells were champed at -80 mV, decrease in the extracellular Na+ resulted in a positive shift of the holding current. In cell-attached voltage-clamp and whole-cell current-clamp configurations, the reduction of extracellular Na+ caused hyperpolarisation. The holding current shifted in negative direction when extracellular K+ concentration was increased from 5 mM to 50 mM in the presence of K+ channel blockers, Ba2+ and TEA, indicating cation nature of persistent conductance. RT-PCR analyses of pars intermedia tissues detected mRNAs of TRPV1, TRPV4, TRPC6, and TRPM3-5. The TRPV channel blocker, ruthenium red, shifted the holding current in positive direction, and significantly decreased the resting [Ca2+]i. These results indicate operation of a constitutive cation conductance sensitive to ruthenium red, which regulates resting membrane potential and [Ca2+]i in rat melanotrophs.
- MeSH
- kationtové kanály TRPV antagonisté a inhibitory metabolismus MeSH
- krysa rodu rattus MeSH
- melanotropní buňky metabolismus MeSH
- metoda terčíkového zámku MeSH
- potkani Wistar MeSH
- rutheniová červeň farmakologie MeSH
- sodík metabolismus MeSH
- vápník metabolismus MeSH
- vápníkové kanály - typ L metabolismus MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Hypotonic solution alters ion channel activity, but little attention has been paid to voltage-dependent sodium channels. The aim of this study was to investigate the effects of hypotonic solution on transient sodium currents (INaT) and persistent sodium currents (INaP). We also explored whether the intracellular signal transduction systems participated in the hypotonic modifications of sodium currents. INaT and INaP were recorded by means of whole-cell patch-clamp technique in isolated rat ventricular myocytes. Our results revealed that hypotonic solution reduced INaT and simultaneously augmented INaP with the occurrence of interconversion between INaT and INaP. Hypotonic solution shifted steady-state inactivation to a more negative potential, prolonged the time of recovery from inactivation, and enhanced intermediate inactivation (IIM). Ruthenium red (RR, inhibitor of TRPV4), bisindolylmaleimide VI (BIM, inhibitor of PKC), Kn-93 (inhibitor of Ca/CaMKII) and BAPTA (Ca2+-chelator) inhibited the effects of hypotonic solution on INaT and INaP. Therefore we conclude that hypotonic solution inhibits INaT, enhances INaP and IIM with the effects being reversible. TRPV4 and intracellular Ca2+, PKC and Ca/CaMKII participate in the hypotonic modifications of sodium currents.
- MeSH
- benzylaminy farmakologie MeSH
- chelátory farmakologie MeSH
- EGTA analogy a deriváty farmakologie MeSH
- financování organizované MeSH
- hypotonické roztoky MeSH
- indoly farmakologie MeSH
- inhibitory proteinkinas farmakologie MeSH
- kardiomyocyty metabolismus účinky léků MeSH
- kationtové kanály TRPV antagonisté a inhibitory metabolismus MeSH
- kinetika MeSH
- krysa rodu rattus MeSH
- maleimidy farmakologie MeSH
- membránové potenciály MeSH
- metoda terčíkového zámku MeSH
- proteinkinasa C antagonisté a inhibitory metabolismus MeSH
- proteinkinasa závislá na vápníku a kalmodulinu typ 2 antagonisté a inhibitory metabolismus MeSH
- rutheniová červeň farmakologie MeSH
- signální transdukce účinky léků MeSH
- sodík metabolismus MeSH
- srdeční komory cytologie metabolismus účinky léků MeSH
- sulfonamidy farmakologie MeSH
- vápník metabolismus MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
