In vitro patch-clamp studies in skin fibroblasts

Goodwin, Paul A.; Campbell, Katherine E.; Evans, Bronwen Alice James; Wann, K. T.

doi:10.1016/s1056-8719(98)00029-x

Cited by 6 publications

(6 citation statements)

References 10 publications

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“…The above value is quite different from those in endothelial cells of the human umbilical cord vein [5] and vascular smooth muscle cells [24] , which evidenced half-maximal activation at 5. The results of our study of the BK Ca channels in human skin fi broblasts were different from those of a previous report by Goodwin et al [25] , which involved the cultured fi broblasts of human external genitalia. Their BK Ca channels exhibited smaller conductance (187 pS) in symmetrical K + solution (140 m M ) than did ours, or those of other reports (200-300 pS) [21,22] .…”

Section: Discussioncontrasting

confidence: 99%

Nitric Oxide Stimulates a Large-Conductance Ca2+-Activated K+ Channel in Human Skin Fibroblasts through Protein Kinase G Pathway

Lim

Yun²,

Kim³

et al. 2005

Skin Pharmacol Physiol

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In order to investigate the large-conductance Ca²⁺-activated K⁺ (BK_Ca) channel and determine the effects of nitric oxide (NO) on the channel in human skin fibroblasts, we performed electrophysiological patch clamp recordings on 5th-passage cells of human genital skin cultures. The whole-cell outward K⁺ current was increased with depolarization, and proved to be sensitive to NS1619 (a selective BK_Ca channel activator) and iberiotoxin (a specific BK_Cachannel inhibitor). The single-channel currents showed 226 pS of mean conductance in symmetrical K⁺. Sodium nitroprusside (SNP; an NO donor) significantly increased the K⁺ current amplitude in the whole-cell mode, and open probability of the channel (NPo) in the cell-attached mode, but not in the inside-out mode. S-nitroso-N-acetylpenicillamine (an NO donor) and 8-Br-cGMP (a membrane-permeant cGMP analogue) also increased the BK_Cachannel activity. The stimulatory effect of SNP on BK_Ca channels was inhibited by pretreatment with 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (a soluble guanylyl cyclase inhibitor), or KT5823 [a specific protein kinase G (PKG) inhibitor]. Cytoplasmic PKG also increased the channel activity in inside-out patches. In conclusion, the present data indicate that BK_Ca channels constitute a significant fraction of K⁺ current in human skin fibroblasts, and that NO increases NPo of BK_Ca channels, which are mediated via the cGMP/PKG pathway, without direct effects on the channel.

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Section: Discussioncontrasting

confidence: 99%

Nitric Oxide Stimulates a Large-Conductance Ca2+-Activated K+ Channel in Human Skin Fibroblasts through Protein Kinase G Pathway

Lim

Yun²,

Kim³

et al. 2005

Skin Pharmacol Physiol

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“…According to our results, chemically-induced MSC from adult rat bone marrow did not present K + or Na + currents (in contrast to what was recorded in immature neurons). Moreover, chemically-induced MSC are electrophysiologically different from fibroblasts, since they present a class of maxi K + channels [54].…”

Section: Discussionmentioning

confidence: 99%

Chemically-Induced RAT Mesenchymal Stem Cells Adopt Molecular Properties of Neuronal-Like Cells but Do Not Have Basic Neuronal Functional Properties

et al. 2009

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Induction of adult rat bone marrow mesenchymal stem cells (MSC) by means of chemical compounds (β-mercaptoethanol, dimethyl sulfoxide and butylated hydroxyanizole) has been proposed to lead to neuronal transdifferentiation, and this protocol has been broadly used by several laboratories worldwide. Only a few hours of MSC chemical induction using this protocol is sufficient for the acquisition of neuronal-like morphology and neuronal protein expression. However, given that cell death is abundant, we hypothesize that, rather than true neuronal differentiation, this particular protocol leads to cellular toxic effects. We confirm that the induced cells with neuronal-like morphology positively stained for NF-200, S100, β-tubulin III, NSE and MAP-2 proteins. However, the morphological and molecular changes after chemical induction are also associated with an increase in the apoptosis of over 50% of the plated cells after 24 h. Moreover, increased intracellular cysteine after treatment indicates an impairment of redox circuitry during chemical induction, and in vitro electrophysiological recordings (patch-clamp) of the chemically induced MSC did not indicate neuronal properties as these cells do not exhibit Na+ or K+ currents and do not fire action potentials. Our findings suggest that a disruption of redox circuitry plays an important role in this specific chemical induction protocol, which might result in cytoskeletal alterations and loss of functional ion-gated channels followed by cell death. Despite the neuronal-like morphology and neural protein expression, induced rat bone marrow MSC do not have basic functional neuronal properties, although it is still plausible that other methods of induction and/or sources of MSC can achieve a successful neuronal differentiation in vitro.

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“…In tissues such as the brain and heart, in which cells fulfill different functional roles, the presence of multiple transcripts would not be surprising. While arteries do have cell types other than smooth muscle cells, neither endothelial cells 30 or fibroblasts 31 have been reported to express Kv channel transcripts. Either all smooth The voltage dependence of K v activation was determined from tail currents measured at Ϫ60 mV after activation from a Ϫ80 mV holding potential to various voltages from Ϫ60 to ϩ60 mV.…”

Section: Discussionmentioning

confidence: 99%

Differential Expression of Voltage-Gated K ⁺ Channel Genes in Arteries From Spontaneously Hypertensive and Wistar-Kyoto Rats

2001

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Abstract-Voltage-gated K ϩ currents play an important role in determining membrane potential, intracellular Ca 2ϩ , and contraction in arterial smooth muscle. In this study, the expression of genes encoding voltage-gated K ϩ channels of the Kv1.X family was compared in arteries from spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). Expression of Kv1.X in thoracic aorta, mesenteric arteries, tail artery, and heart was determined, both qualitatively and quantitatively, by reverse transcription-polymerase chain reaction. Our results demonstrate distinct but overlapping patterns of expression in vascular tissues. In general, Kv1.2 and Kv1.5 were most highly represented, and the levels of Kv1.2 were significantly larger in all tissues from SHR. Levels of Kv1.5 in arteries did not differ significantly between strains but were greater in SHR heart. Moderate levels of Kv1.3 and Kv␤1.1 expression were also found in all tissues and were larger in SHR. Kv1.1 expression was not different between the 2 strains, and no significant expression of Kv1.4 (except in heart and aorta), Kv1.6, or Kv␤2.1 was observed in either strain. Kv1.2 and Kv1.5 transcripts represent Ϸ1 to 2 parts/10 5 of total mesenteric arterial RNA with Ϸ2-to 5-fold lower levels in aorta and tail artery. Whole-cell voltage-gated K ϩ channel currents, recorded from mesenteric arterial myocytes, were larger in SHR than WKY (eg, at 0 mV: 7.3Ϯ0.8 versus 10.9Ϯ1.2 pA/pF). The voltage dependence of activation was more negative in SHR (V 0.5 : Ϫ20Ϯ4 mV versus Ϫ32Ϯ3 mV) but that of availability was not different. These results indicate that Kv1.X genes are differentially expressed between WKY and SHR (especially Kv1.2 and Kv␤1.1). These differences in gene expression are associated with a greater voltage-gated K ϩ channel current density in SHR and shifted voltage-dependent activation compared with WKY. These differences may be a compensatory mechanism related to the membrane potential depolarization in SHR or some manifestation thereof.

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In vitro patch-clamp studies in skin fibroblasts

Cited by 6 publications

References 10 publications

Nitric Oxide Stimulates a Large-Conductance Ca<sup>2+</sup>-Activated K<sup>+</sup> Channel in Human Skin Fibroblasts through Protein Kinase G Pathway

Nitric Oxide Stimulates a Large-Conductance Ca<sup>2+</sup>-Activated K<sup>+</sup> Channel in Human Skin Fibroblasts through Protein Kinase G Pathway

Chemically-Induced RAT Mesenchymal Stem Cells Adopt Molecular Properties of Neuronal-Like Cells but Do Not Have Basic Neuronal Functional Properties

Differential Expression of Voltage-Gated K ⁺ Channel Genes in Arteries From Spontaneously Hypertensive and Wistar-Kyoto Rats

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In vitro patch-clamp studies in skin fibroblasts

Cited by 6 publications

References 10 publications

Nitric Oxide Stimulates a Large-Conductance Ca<sup>2+</sup>-Activated K<sup>+</sup> Channel in Human Skin Fibroblasts through Protein Kinase G Pathway

Nitric Oxide Stimulates a Large-Conductance Ca<sup>2+</sup>-Activated K<sup>+</sup> Channel in Human Skin Fibroblasts through Protein Kinase G Pathway

Chemically-Induced RAT Mesenchymal Stem Cells Adopt Molecular Properties of Neuronal-Like Cells but Do Not Have Basic Neuronal Functional Properties

Differential Expression of Voltage-Gated K + Channel Genes in Arteries From Spontaneously Hypertensive and Wistar-Kyoto Rats

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Differential Expression of Voltage-Gated K ⁺ Channel Genes in Arteries From Spontaneously Hypertensive and Wistar-Kyoto Rats