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Curcumin (1,7-bis(4-hydroxy-3-methoxyphenyl)-1E,6E-heptadiene-3,5-dione or diferuloyl methane) is a polyphenol derived from the Curcuma longa plant, commonly known as turmeric. This substance has been used extensively in Ayurvedic medicine for centuries for its anti-oxidant, analgesic, anti-inflammatory and antiseptic activity. More recently curcumin has been found to possess anti-cancer properties linked to its pro-apoptotic and anti-proliferative actions. The underlying mechanisms of these diverse effects are complex, not fully elucidated and subject of intense scientific debate. Despite increasing evidence indicating that different cation channels can be a molecular target for curcumin, very little is known about the effect of curcumin on chloride channels. Since, (i) the molecular structure of curcumin indicates that the substance could potentially interact with chloride channels, (ii) chloride channels play a role during the apoptotic process and regulation of the cell volume, and (iii) apoptosis is a well known effect of curcumin, we set out to investigate whether or not curcumin could (i) exert a modulatory effect (direct or indirect) on the swelling activated chloride current ICl(swell) in a human cell system, therefore (ii) affect cell volume regulation and (iii) ultimately modulate cell survival. The ICl(swell) channels, which are essential for regulating the cell volume after swelling, are also known to be activated under isotonic conditions as an early event in the apoptotic process. Here we show that long-term exposure of a human kidney cell line to extracellular 0.1-10 μM curcumin modulates ICl(swell) in a dose-dependent manner (0.1 μM curcumin is ineffective, 0.5-5.0 μM curcumin increase, while 10 μM curcumin decrease the current), and short-term exposure to micromolar concentrations of curcumin does not affect ICl(swell) neither if applied from the extracellular nor from the intracellular side - therefore, a direct effect of curcumin on ICl(swell) can be ruled out. Furthermore, we show that curcumin exposure induces apoptosis in human kidney cells, and at a concentration of 5.0-10 μM induces the appearance of a sub-population of cells with a dramatically increased volume. In these cells the regulation of the cell volume seems to be impaired, most likely as a consequence of the ICl(swell) blockade. Similarly, 50 μM curcumin induced apoptosis, caused cell cycle arrest in G1-phase and increased the volume of human colorectal adenocarcinoma HT-29 cells. The cell cycle arrest in G1 phase may be the mechanism underlying the volume increase observed in this cell line after exposure to curcumin., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Because oxidative stress is a component of gastrointestinal injury, we investigated the effect of H(2)O(2) on transintestinal transport using isolated rat jejunum incubated in vitro. Millimolar concentrations of H(2)O(2) inhibited all the tested parameters without inducing any cytotoxic effect. Electrophysiological experiments indicated that H(2)O(2) decreases significantly both short circuit current and transepithelial electrical potential difference without affecting transepithelial resistance. The possibility that H(2)O(2) could influence (Na+, K+) -ATPase activity was explored using isolated basolateral membranes. Besides H(2)O(2), free radicals (O(2)(*-), HO*) were generated using different iron-dependent and independent systems; (Na+, K+) -ATPase activity was inhibited after membrane exposure to all ROS tested. The inhibition was prevented by allopurinol, superoxide dismutase or desferrioxamine. Western blot analysis showed a decreased expression of the alpha(1)-subunit of (Na+, K+) -ATPase. We conclude that H(2)O(2) may be a modulator of jejunal ion and water transport by multiple mechanisms, among which a significant inhibition of the basolateral (Na+, K+) -ATPase.

In guinea pig gallbladder epithelial cells, an increase in intracellular cAMP levels elicits the rise of anion channel activity. We investigated by patch-clamp techniques whether K(+) channels were also activated. In a cell-attached configuration and in the presence of theophylline and forskolin or 8-Br-cAMP in the cellular incubation bath, an increase of the open probability (P(o)) values for Ca(2+)-activated K(+) channels with a single-channel conductance of about 160 pS, for inward current, was observed. The increase in P(o) of these channels was also seen in an inside-out configuration and in the presence of PKA, ATP, and cAMP, but not with cAMP alone; phosphorylation did not influence single-channel conductance. In the inside-out configuration, the opioid loperamide (10(-5) M) was able to reduce P(o) when it was present either in the microelectrode filling solution or on the cytoplasmic side. Detection in the epithelial cells by RT-PCR of the mRNA corresponding to the alpha subunit of large-conductance Ca(2+)-activated K(+) channels (BK(Ca)) indicates that this gallbladder channel could belong to the BK family. Immunohistochemistry experiments confirm that these cells express the BK alpha subunit, which is located on the apical membrane. Other K(+) channels with lower conductance (40 pS) were not activated either by 8-Br-cAMP (cell-attached) or by PKA + ATP + cAMP (inside-out). These channels were insensitive to TEA(+) and loperamide. The data demonstrate that under conditions that induce secretion, phosphorylation activates anion channels as well as Ca(2+)-dependent, loperamide-sensitive K(+) channels present on the apical membrane., (©2002 Elsevier Science (USA).)

The effect of follicle-stimulating hormone (FSH) on the electrical properties of TM4-Sertoli cells was investigated. Addition of 5 IU/ml FSH caused a dose-dependent and reversible depolarization of the resting membrane potential by +15.3 +/- 1.0 mV accompanied by a decrease of the input resistance. The depolarization was completely abolished in chloride-free solutions. The reversal potential of the effect was close to the calculated reversal potential for chloride. We conclude that FSH activates a chloride conductance in cultured TM4 Sertoli cells.

The MDCK cell has proven to be a useful model cell line for the study of properties and regulation of renal epithelial ion channels. Patch clamp studies disclosed the existence of several K+ channels and of a Cl- channel, and their regulation by hormones, cell volume, trace elements and drugs. Most hormones affect K+ channels at least in part by increasing cytosolic Ca2+. However, indirect evidence points to additional mechanisms contributing to K+ channel activation. Cell swelling activates both K+ channels and unselective anion channels. ICln, a protein cloned from MDCK cells, is either a Cl- channel or a regulator of thereof. ICln is up-regulated by cellular acidification and is crucial for rapid regulatory cell volume decrease.

The kidney is a main target organ for cadmium toxicity. The present study has been performed to test for effects of cadmium on electrical properties of cultured subconfluent kidney (MDCK) cells. Cadmium leads to a rapid, sustained and reversible hyperpolarization of the cell membrane, paralleled by an increase of the potassium selectivity and a decrease of the resistance. Thus, cadmium increases the potassium conductance of the cell membrane. The half maximal effect is elicited congruent to 0.2 microM, a concentration encountered during chronic cadmium intoxication. At extracellular calcium concentration reduced to less than 0.1 microM, 5 microM cadmium leads to a transient hyperpolarization, which can be elicited only once. High concentrations (50 microM) of cadmium lead to a sustained hyperpolarization even at extracellular calcium concentrations of less than 0.1 microM. According to fluorescence measurements cadmium leads to an increase of intracellular calcium activity, which is sustained at 1 mM and transient at less than 1 microM extracellular calcium activity. In conclusion, cadmium at low concentrations enhances the potassium conductance in a calcium dependent way. The observations suggest that cadmium enhances intracellular calcium both by recruitment from intracellular stores and by modification of calcium transport across the cell membrane. At high concentrations cadmium enhances the potassium conductance independently from enhanced intracellular calcium activity.

Fura2 - fluorescence was utilized to test for the effect of extracellular nucleotides on intracellular calcium concentration of subconfluent Madin-Darby Canine Kidney (MDCK)-cells. Extracellular ATP (10 mumol/l) and UTP (10 mumol/l) lead to rapid (within seconds), sustained, and fully reversible enhancement of intracellular calcium concentration from 138 +/- 9 nmol/l (n = 27), to 1561 +/- 260 nmol/l (n = 10) and 3435 +/- 949 nmol/l (n = 5), respectively. Half maximal effects are observed at some 1 mumol/l. In the absence of extracellular calcium the effect of ATP is transient, pointing to release of intracellular calcium. The sustained effect in the presence of extracellular calcium indicates that the nucleotides in addition recruit calcium from extracellular space.