Gramicidin-Perforated Patch Analysis on HCO 3 − Secretion Through a Forskolin-Activated Anion Channel in Rat Parotid Intralobular Duct Cells

Hirono, Chikara; Nakamoto, Tetsuji; Sugita, Makoto; Iwasa, Yoshiko; Akagawa, Yasumasa; Shiba, Yoshiki

doi:10.1007/s002320010054

Cited by 21 publications

(22 citation statements)

References 27 publications

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“…Our findings thus accord with the results of earlier, more detailed studies of cells isolated from adult animals that identified CFTR-like anion channels in distal lung epithelial cells (16,29). However, in our hands, the cAMP-evoked increase in G Cl Ϫ consisted of an initial peak followed by a slower, more sustained phase, whereas previous studies of anion currents flowing through CFTR-like channels have reported that cAMP causes a much more sustained response (12,16). The reason for this discrepancy between these two studies is unknown but may reflect the different origin of the cells used in the two studies.…”

Section: Discussionsupporting

confidence: 92%

β-Adrenoceptor-mediated control of apical membrane conductive properties in fetal distal lung epithelia

Collett

Ramminger

Olver

et al. 2002

American Journal of Physiology-Lung Cellular and Molecular Phys

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Distal lung epithelial cells isolated from fetal rats were cultured (48 h) on permeable supports so that transepithelial ion transport could be quantified electrometrically. Unstimulated cells generated a short-circuit current (Isc) that was inhibited (ϳ80%) by apical amiloride. The current is thus due, predominantly, to the absorption of Na ϩ from the apical solution. Isoprenaline increased the amiloride-sensitive Isc about twofold. Experiments in which apical membrane Na ϩ currents were monitored in basolaterally permeabilized cells showed that this was accompanied by a rise in apical Na ϩ conductance (GNaϩ). Isoprenaline also increased apical Cl Ϫ conductance (GClϪ) by activating an anion channel species sensitive to glibenclamide but unaffected by 4,4Ј-diisothiocyanatostilbene-2,2Ј-disulfonic acid (DIDS). The isoprenaline-evoked changes in G Na ϩ and GClϪ could account for the changes in Isc observed in intact cells. Glibenclamide had no effect upon the isoprenaline-evoked stimulation of I sc or GNaϩ demonstrating that the rise in GClϪ is not essential to the stimulation of Na ϩ transport. alveolar ion transport; Ussing chambers; permeabilized epithelia THROUGHOUT FETAL LIFE the distal lung epithelia secrete fluid into the developing air spaces (31) and establish a distending pressure that is crucial to lung morphogenesis (11). However, this liquid must be removed from the lungs if the newborn infant is to breathe at birth. The absorption of this liquid occurs during the final stages of gestation and is dependent upon the active withdrawal of Na ϩ from the lung lumen, a process that can be controlled via ␤-adrenoceptors (23,30,(36)(37)(38). The means by which this control is achieved are not fully understood, but there is evidence that the process involves a rise in apical Na ϩ conductance (G Na ϩ) (15,20). It is also known that ␤-adrenoceptor agonists can increase apical Cl Ϫ conductance (G Cl Ϫ) in these cells, and it has been suggested that this may facilitate Na ϩ transport by increasing the driving force for Na ϩ entry (16,29). It is, however, difficult to see how the ionic gradients that normally prevail in epithelial cells could allow Na ϩ transport to be controlled in this way (see Refs. 20,44). To clarify the means by which ␤-adrenoceptor agonists can control Na ϩ absorption in the distal lung epithelia, we now explore the effects of isoprenaline upon the conductive properties of the apical membrane. METHODS Isolation and culture of rat fetal distal lung epithelial cells.Fetuses removed from anesthetized (3% halothane), 20-day pregnant (term ϭ 22 days) rats were immediately decapitated, and their lung tissue was collected into ice-cold, Ca 2ϩ -and Mg 2ϩ -free Hanks' balanced salt solution. The anesthetized animals were then killed (cervical dislocation/exsanguination) before regaining consciousness. The fetal lung tissue was chopped into pieces (Ͻ0.5 mm) and disaggregated using 0.2% trypsin/0.012% DNase (2 ϫ 20 min, 37°C) followed by 0.1% collagenase/0.012% DNase [15 min, 37°C, both in Dulbecco's mo...

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Section: Discussionsupporting

confidence: 92%

β-Adrenoceptor-mediated control of apical membrane conductive properties in fetal distal lung epithelia

Collett

Ramminger

Olver

et al. 2002

American Journal of Physiology-Lung Cellular and Molecular Phys

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“…The details have already been described in a previous report [18]. Briefly, gramicidin D (Sigma) was dissolved in methanol (10 mg/ml) and diluted 100 times with a standard KCl-rich pipette solution.…”

Section: Membrane Potential Measurementsmentioning

confidence: 99%

“…The membrane potential of submandibular acinar cells was measured using a whole cell patch-clamp technique [18]. The resting membrane potential was approximately -40 to -50 mV.…”

Section: Membrane Potential Measurementmentioning

confidence: 99%

Membrane potential modulation of ionomycin-stimulated Ca2+ entry via Ca2+/H+ exchange and SOC in rat submandibular acinar cells

et al. 2010

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Ionomycin (IM) at 5 microM mediates the Ca(2+)/H(+) exchange, while IM at 1 microM activates the store-operated Ca(2+) entry channels (SOCs). In this study, the effects of depolarization on both pathways were examined in rat submandibular acinar cells by increasing extracellular K(+) concentration ([K(+)](o)). IM (5 microM, the Ca(2+)/H(+) exchange) increased the intracellular Ca(2+) concentration ([Ca(2+)](i)) to an extremely high value at 151 mM [K(+)](o). However, with increasing [K(+)](o), the rates of Ca(2+) entry decreased in a linear relationship. The reversal potential (E (rev)) for the Ca(2+)/H(+) exchange was +93 mV, suggesting that IM (5 microM) exchanges 1 Ca(2+) for 1 H(+). Thus, depolarization decreases the Ca(2+) influx via the Ca(2+)/H(+) exchange because of its electrogenicity (1 Ca(2+) for 1 H(+)). On the other hand, IM (1 microM, the SOCs) abolished an increase in [Ca(2+)](i) at 151 mM [K(+)](o). With increasing [K(+)](o), the rate of Ca(2+) entry immediately decreased linearly. The E (rev) for the SOC was +3.7 mV, suggesting that the SOCs are nonselective cation channels and less selective for Ca(2+) over Na(+) (P (Ca)/P (Na) = 8.2). Moreover, an increase in extracellular Ca(2+) concentration (20 mM) enhanced the Ca(2+) entry via the SOCs at 151 mM [K(+)](o), suggesting depolarization does not inhibit the SOCs and decreases the driving force for the Ca(2+) entry. This suggests that membrane potential changes induced by a secretory stimulation finely regulate the [Ca(2+)](i) via the SOCs in rat submandibular acinar cells. In conclusion, IM increases [Ca(2+)](i) via two pathways depending on its concentration, the exchange of 1 Ca(2+) for 1 H(+) at 5 muM and the SOCs at 1 microM.

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“…Cl − secretion can be measured as Cl − secretory currents by using the gramicidin-perforated patch-clamp technique (14,15,32,34 -activated Cl − currents. A patch/whole-cell clamp amplifier (CEZ-2400; Nihon Kohden, Tokyo, Japan) was used to measure whole-cell currents.…”

Section: +mentioning

confidence: 99%

“…All measurements of the oscillatory Cl − secretory current were performed with the gramicidin-perforated patch-recording method (14,15,32,34). A pipette solution containing 150 mM KCl, 10 mM HEPES, and 0.1 mg/mL gramicidin D (Sigma-Aldrich) was adjusted to pH 7.4 using KOH.…”

Section: Measurement Of the Oscillatory CLmentioning

confidence: 99%

Involvement of cytoskeletal integrity in the regulation of Cl- and amylase secretion from rat parotid acinar cells

et al. 2008

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The cytoskeleton serves as a signal modulator for Ca 2+ and cAMP-regulated cell functions including the secretion of ions and granule contents. The interaction between Ca 2+ and cAMP signaling systems potentiates amylase secretion and suppresses Cl − secretion in the parotid glands. In this study, we investigated the role of the cytoskeleton in the modulation of Cl − and amylase secretion from rat parotid acinar cells upon activation of each intracellular signaling system and their interaction. Cytochalasin D markedly inhibited the Ca 2+ -activated outwardly rectifying Cl − current at positive membrane potentials and carbachol (CCh)-induced Cl − currents in the whole-cell configuration at −80 mV, whereas colchicine enhanced Cl − currents. Cytochalasin D, but not colchicine, markedly inhibited CCh-induced Cl − secretion. Synergistic actions of CCh and forskolin on Cl − and amylase secretion were observed even in the presence of cytochalasin D. These results suggest that the synergistic effects of Ca 2+ and cAMP signaling systems on amylase and Cl − secretion do not require actin filament integrity but that secretion by the two signals themselves does require actin filament integrity.Actin and tubulin cytoskeletons are involved in the regulation of cell morphology and intracellular transport. They also affect the membrane permeability and secretory functions of glandular cells; cytoskeletal integrity modulates amylase secretion by β-adrenergic agonists (4,5,24,37) and cytoskeletal organization is related to the regulation of ion channels (1, 12), exchangers (16) and transporters (7) involved in the secretion of salivary fluid during cholinergic stimulation (22,27). Cytoskeletal integrity might regulate Cl −

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Gramicidin-Perforated Patch Analysis on HCO 3 − Secretion Through a Forskolin-Activated Anion Channel in Rat Parotid Intralobular Duct Cells

Cited by 21 publications

References 27 publications

β-Adrenoceptor-mediated control of apical membrane conductive properties in fetal distal lung epithelia

β-Adrenoceptor-mediated control of apical membrane conductive properties in fetal distal lung epithelia

Membrane potential modulation of ionomycin-stimulated Ca2+ entry via Ca2+/H+ exchange and SOC in rat submandibular acinar cells

Involvement of cytoskeletal integrity in the regulation of Cl- and amylase secretion from rat parotid acinar cells

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