cis‐Fatty acids, which activate protein kinase C, attenuate Na+ and Ca2+ currents in mouse neuroblastoma cells.

D. J. Linden, A. Routtenberg

Research output: Contribution to journalArticlepeer-review

Abstract

1. Activation of protein kinase C (PKC) by phorbol esters or diacylglycerols has been shown to modulate a number of ionic currents carried by Ca2+, K+ and Cl‐. Recently, it has been demonstrated that PKC may be activated by cis‐fatty acids in the absence of either phospholipid or Ca2+. We wished to determine if this new class of PKC‐activating compound would also modulate ionic currents. To this end we applied the whole‐cell voltage‐clamp technique to N1E‐115 neuroblastoma cells. 2. Analysis of families of currents evoked under voltage clamp by depolarizing steps from a holding potential of ‐85 mV during external application of 5 microM‐oleate (a cis‐fatty acid) showed a 36% reduction of the peak inward current with no shift in either the peak or the reversal potential of the current‐voltage relation and no alteration of outward current. 3. External application of the cis‐fatty acids oleate, linoleate and linolenate reversibly attenuated voltage‐dependent Na+ current with approximate half‐maximal dose values of 2, 3, and 10 microM respectively. Oleate was approximately 2 times more potent when applied internally (ED50 = 1 microM). Externally applied elaidate (a trans‐isomer of oleate) and stearate (a saturated fatty acid) which do not activate PKC, had no effect. Since cis‐fatty acids are known to fluidize membranes, as well as to activate PKC, we sought to dissociate these functions by applying compounds that fluidize membranes but do not activate PKC: methyloleate and lysophosphatidylcholine. Neither compound affected Na+ current when applied externally at concentrations of 1‐50 microM. 4. In contrast to cis‐fatty acids, three classical PKC activators, phorbol‐12.13‐dibutyrate (PDB), phorbol‐12.13‐diacetate (PDA), and 1.2‐oleoylacetylglycerol (OAG) were found to have no effect on the voltage‐dependent Na+ current when applied externally at 10 nM‐1 microM (phorbol esters) or 1‐150 microM (OAG) for incubation periods up to 1 h. 5. External application of the PKC inhibitors polymyxin B, H‐7, sphingosine and staurosporine blocked the attenuation of the Na+ current by cis‐fatty acid in a dose‐dependent manner, with maximal inhibition occurring at doses of 50, 10, 200 and 0.1 microM, respectively. The cyclic nucleotide‐dependent protein kinase inhibitor H‐8 was much less effective in blocking the cis‐fatty acid effect. Polymyxin B and staurosporine were more potent when applied internally. 6. Chronic (24 h) exposure to 1 microM phorbol‐12‐myristate‐13‐acetate (TPA) was employed to down‐regulate PKC.(ABSTRACT TRUNCATED AT 400 WORDS)

Original languageEnglish (US)
Pages (from-to)95-119
Number of pages25
JournalThe Journal of physiology
Volume419
Issue number1
DOIs
StatePublished - Dec 1 1989
Externally publishedYes

ASJC Scopus subject areas

  • Physiology

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