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  • 1
    Electronic Resource
    Electronic Resource
    Expression of amiloride-sensitive Na^+ channels of hen lower intestine in Xenopus oocytes: Electrophysiological studies on the dependence of varying NaCl intake
    Amsterdam : Elsevier
    Biochimica et Biophysica Acta (BBA)/Biomembranes 1111 (1992), S. 159-164 
    Details
    ISSN: 0005-2736
    Keywords: (Colon) ; (Coprodeum) ; (Hen intestine) ; Amiloride ; Current-voltage relationship ; Expression ; Oocyte ; Sodium ion channel
    Source: Elsevier Journal Backfiles on ScienceDirect 1907 - 2002
    Topics: Biology , Chemistry and Pharmacology , Medicine , Physics
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1016/0005-2736(92)90306-7
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  • 2
    Electronic Resource
    Electronic Resource
    Cystic fibrosis and non-cystic-fibrosis human nasal epithelium show analogous Na+ absorption and reversible block by phenamil (1997)
    Springer
    Pflügers Archiv 434 (1997), S. 19-24 
    Details
    ISSN: 1432-2013
    Keywords: Key words Cystic fibrosis ; Human nasal epithelium ; Primary culture ; Epithelial Na+ channel ; Amiloride ; Phenamil
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  Transepithelial short-circuit current (I SC), potential (V T) and resistance (R T) of confluent monolayers of human nasal epithelium cultured from patients with and without cystic fibrosis (CF) were measured. In our Ussing chamber experiments with monolayers derived from non-CF and CF patients neither I SC (non-CF: 14.1 ± 1.0 μA/cm2, n = 77; CF: 16.7 ± 1.5 μA/cm2, n = 42), nor R T (non-CF: 288 ± 15 Ω· cm2; CF: 325 ± 20 Ω· cm2) showed any significant differences, only V T showed moderate but significant different values (non-CF: –3.6 ± 0.4 mV; CF: –5.6 ± 0.7 mV, respectively). Total I SC in CF cells was nearly completely inhibited by amiloride (92 ± 9.6%), while in non-CF tissue amiloride-insensitive conductances mediated a considerable amount of the I SC (36.3 ± 6.1%), indicating a lower activity of amiloride-sensitive Na+ conductances in non-CF cells. In both tissues the amiloride-sensitive I SC could also be blocked by the amiloride analogues benzamil, phenamil and 5-(N-ethyl-N-isopropyl)2’,4’-amiloride (EIPA) with different affinities. However, amiloride had a significant lower affinity in CF tissue (half-maximal blocker concentration, K 1/2 = 586 ± 59 nM) compared with non-CF tissue (K 1/2 = 294 ± 22 nM). Astonishingly, phenamil, a blocker which irreversibly blocks all epithelial Na+ channels hitherto described, inhibited the Na+ conductances of human nasal epithelium in a completely reversible way, but nevertheless with high affinity (non-CF: K 1/2 = 12.5 ± 1.2 nM; CF: K 1/2 = 17.1 ± 1.1 nM). Even in high doses none of these blockers had any effect on intracellular Ca2+ concentration as measured with Fura-2. From these findings, we conclude that the epithelial Na+ conductances of human CF nasal epithelium show modified regulation or are functionally different from those of other tissues.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050358
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  • 3
    Electronic Resource
    Electronic Resource
    cAMP-activation of amiloride-sensitive Na+ channels from guinea-pig colon expressed in Xenopus oocytes (1996)
    Springer
    Pflügers Archiv 431 (1996), S. 913-922 
    Details
    ISSN: 1432-2013
    Keywords: Key wordsXenopus oocytes ; Guinea-pig colon ; Na+ channel ; Single-channel analysis ; Amiloride ; Benzamil ; cAMP
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Guinea-pig distal colonic mRNA injection into Xenopus laevis oocytes resulted in expression of functional active epithelial Na+ channels in the oocyte plasma membrane. Poly(A)+ RNA was extracted from distal colonic mucosa of animals fed either a high-salt (HS) or a low-salt (LS) diet. The electrophysiological properties of the expressed amiloride-sensitive Na+ conductances were investigated by conventional two-electrode voltage-clamp and patch-clamp measurements. Injection of poly(A)+ RNA from HS-fed animals [from hereon referred to as HS-poly(A)+ RNA] into oocytes induced the expression of amiloride-sensitive Na+ conductances. On the other hand, oocytes injected with poly(A)+ RNA from LS-fed animals [LS-poly(A)+ RNA] expressed a markedly larger amount of amiloride-blockable Na+ conductances. LS-poly(A)+ RNA-induced conductances were completely inhibitable by amiloride with a K i of 77 nM, and were also blocked by benzamil with a K i of 1.8 nM. 5-(N-Ethyl-N-isopropyl)-amiloride (EIPA), even in high doses (25 μM), had no detectable effect on the Na+ conductances. Expressed amiloride-sensitive Na+ channels could be further activated by cAMP leading to nearly doubled clamp currents. When Na+ was replaced by K+, amiloride (1 μM) showed no effect on the clamp current. Single-channel analysis revealed slow gating behaviour, open probabilities (P o) between 0.4 and 0.9, and slope conductances of 3.8 pS for Na+ and 5.6 pS for Li+. The expressed channels showed to be highly selective for Na+ over K+ with a permeability ratio P Na/P K 〉 20. Amiloride (500 nM) reduced channel P o to values 〈 0.05. All these features make the guinea-pig distal colon of LS-fed animals an interesting mRNA source for the expression of highly amiloride-sensitive Na+ channels in Xenopus oocytes, which could provide new insights in the regulatory mechanism of these channels.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004240050085
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  • 4
    Electronic Resource
    Electronic Resource
    Ion transport across leech integument (1993)
    Springer
    Journal of comparative physiology 163 (1993), S. 153-159 
    Details
    ISSN: 1432-136X
    Keywords: Na+ channels ; Amiloride ; Benzamil ; Noise analysis ; Leech, Hirudo medicinalis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The dorsal skin of the leech Hirudo medicinalis was used for electrophysiological measurements performed in Ussing chambers. The leech skin is a tight epithelium (transepithelial resistance = 10.5±0.5 kΩ· cm-2) with an initial short-circuit current of 29.0±2.9 μA·cm-2. Removal of Na+ from the apical bath medium reduced short-circuit current about 55%. Ouabain (50μmol·l-1) added to the basolateral solution, depressed the short-circuit current completely. The Na+ current saturated at a concentration of 90 mmol Na+·l-1 in the apical solution (K M=11.2±1.8 mmol·l-1). Amiloride (100 μmol·l-1) on the apical side inhibited ca. 40% of the Na+ current and indicated the presence of Na+ channels. The dependence of Na+ current on the amiloride concentration followed Michaclis-Menten kinetics (K i=2.9±0.4 μmol·l-1). The amiloride analogue benzamil had a higher affinity to the Na+ channel (K i=0.7±0.2 μmol·l-1). Thus, Na+ channels in leech integument are less sensitive to amiloride than channels known from vertebrate epithelia. With 20 mmol Na+·l-1 in the mucosal solution the tissue showed an optimum amiloride-inhibitable current, and the amiloride-sensitive current under this condition was 86.8±2.3% of total short-circuit current. Higher Na+ concentrations lead to a decrease in amiloride-blockade short-circuit current. Sitmulation of the tissue with cyclic adenosine monophosphate (100 μmol·l-1) and isobutylmethylxanthine (1 mmol·l-1) nearly doubled short-circuit current and increased amiloride-sensitive Na+ currents by 50%. By current fluctuation analysis we estimated single Na+ channel current (2.7±0.9 pA) and Na+ channel density (3.6±0.6 channels·μm-2) under control conditions. After cyclic adenosine monophosphate stimulation Na+ channel density increased to 5.4±1.1 channels·μm-2, whereas single Na+ channel current showed no significant change (1.9±0.2 pA). These data present a detailed investigation of an invertebrate epithelial Na+ channel, and show the similarities and differences to vertebrate Na+ channels. Whereas the channel properties are different from the classical vertebrate Na+ channel, the regulation by cyclic adenosine monophosphate seems similar. Stimulation of Na+ uptake by cyclic adenosine monophosphate is mediated by an increasing number of Na+ channels.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00263601
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