Search PubMed⌕ Search

Biomedical subjects

M Paulmichl

Publications and source records attributed to M Paulmichl.

At least 55 records · Page 3Linked to original sources
PubMed1991

Effect of trifluoperazine on renal epithelioid Madin-Darby canine kidney cells.

Following exposure to a number of hormones, the cell membrane in Madin-Darby Canine Kidney (MDCK) cells is hyperpolarized by increase of intracellular calcium activity. The present study has been performed to elucidate the possible role of calmodulin in the regulation of intracellular calcium activity and cell membrane potential. To this end trifluoperazine has been added during continuous recording of cell membrane potential or intracellular calcium. Trifluoperazine leads to a transient increase of intracellular calcium as well as a sustained hyperpolarization of the cell membrane by activation of calcium sensitive K+ channels. Half-maximal effects are observed between 1 and 10 mumol/L trifluoperazine. A further calmodulin antagonist, chlorpromazine, (50 mumol/L), similarly hyperpolarizes the cell membrane. The effects of trifluoperazine are virtually abolished in the absence of extracellular calcium. Pretreatment of the cells with either pertussis toxin or phorbol-ester TPA does not interfere with the hyperpolarizing effect of trifluoperazine. In conclusion, calmodulin is apparently involved in the regulation of calcium transfer across the cell membrane but not in the stimulation of K+ channels by intracellular calcium.

Animals↗
PubMed1991

Further characterization of volume regulatory decrease in cultured renal epitheloid (MDCK) cells.

In Madin Darby canine kidney (MDCK) cells volume regulatory decrease (VRD) is paralleled by a variable, transient hyperpolarization followed by a sustained depolarization of the cell membrane. In the depolarized cells, the cell membrane selectivity is decreased for potassium and increased for chloride. Without knowledge of the cell membrane resistance (Rm), these changes of cell membrane selectivity cannot be translated into conductances, i.e. the observed alterations of ion selectivity could have been due to inhibition of potassium conductance or activation of anion conductance. In the present study Rm has been determined by cellular cable analysis. To this end, three microelectrodes were impaled into three different cells of a cell cluster, current (up to 3 nA) was injected into one cell and the corresponding voltage deflections determined in the other two cells. As a result, exposure of the cells to hypotonic perfusates leads to a marked, sustained reduction of Rm. In the absence of chloride and in the absence of bicarbonate and chloride, the decrease of Rm is only transient. The data indicate that cell swelling leads to a transient increase of potassium conductance followed by a sustained increase of anion conductance. As evident from BCECF fluorescence, exposure of MDCK cells to hypotonic perfusates leads to a significant decrease of intracellular pH, which may in part be due to loss of bicarbonate through the anion conductive pathway.

Animals↗
PubMed1991

Further analysis of ATP-mediated activation of K+ channels in renal epithelioid Madin Darby canine kidney (MDCK) cells.

ATP activates K+ channels by increasing intracellular calcium activity in Madin Darby canine kidney (MDCK) cells. The present study has been performed to test for the involvement of G-proteins and of protein kinase C in the intracellular transmission of these effects. To this end, the effect of ATP on intracellular calcium and K+ channel activity has been studied in cells pretreated with the phorbol ester 12-O-tetradecanoyl-phorbol 13-acetate (TPA) and/or pertussis toxin. The ATP-induced increase of intracellular calcium is not significantly affected by pretreatment with pertussis toxin, is significantly blunted by pretreatment with TPA and is abolished by pretreatment with both pertussis toxin and the phorbol ester. The ATP activation of K+ channels is similarly blunted by pretreatment with TPA, but is not abolished by pretreatment with both the phorbol ester and pertussis toxin. Furthermore, the ATP-induced hyperpolarization is not abolished in cells pretreated with both pertussis toxin and TPA. In those cells, ATP may activate K+ channels by calcium-dependent mechanisms or lead to localized increases of intracellular calcium sufficient to activate the K+ channels but escaping detection with fura-2 fluorescence.

Adenosine Triphosphate↗
PubMed1990

Cobalt activates potassium conductance in the plasma membrane of cultured renal epithelioid (MDCK)-cells.

Cobalt has been shown to stimulate sodium transport across the distal nephron of the newt kidney. The mechanism of this action remained elusive. The present study has been performed to test for effects of cobalt on electrical properties of cultured subconfluent kidney (MDCK)-cells: cobalt (10 microM) 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, cobalt increases the potassium conductance of the cell membrane. The half-maximal effect is elicited by approx. 1 microM. At extracellular calcium concentration reduced to less than 0.1 microM, cobalt (10 microM) leads to a transient hyperpolarization, which can be elicited only once. Thus, cobalt enhances the potassium conductance in a calcium dependent way. At higher concentrations (100 microM) cobalt hyperpolarizes the cell membrane only transiently even in the presence of extracellular calcium. Furthermore 100 microM cobalt interferes with ATP-induced hyperpolarization, which is known to result from calcium mediated activation of K+ channels. Thus, 100 microM cobalt may inhibit ATP-stimulated calcium entry into the cell.

Adenosine Triphosphate↗
PubMed1990

Cadmium enhances potassium conductance in cultured renal epitheloid (MDCK) cells.

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.

Animals↗
PubMed1989

The effect of hypoosmolarity on the electrical properties of Madin Darby canine kidney cells.

The present study has been performed to test for the effect of hypotonic extracellular fluid on the electrical properties of Madin Darby canine kidney (MDCK)-cells. The volume of suspended MDCK-cells is 1,892 +/- 16 fl (n = 8) in isotonic (298.7 mosmol/l) extracellular fluid. Exposure of the cells to hypotonic (230.7 mosmol/l) extracellular fluid is followed by cellular swelling to 2,269 +/- 18 fl (n = 4) and subsequent volume regulatory decrease to 2,052 +/- 22 fl (n = 4) within 512 s. Volume regulatory decrease is abolished by quinidine (1 mmol/l) and by lipoxygenase inhibitor nordihydroguaiaretic acid (50 mumol/l). The potential difference across the cell membrane averages -53.6 +/- 0.9 mV (n = 49) in isotonic extracellular perfusates. Reduction of extracellular osmolarity depolarizes the cell membrane by +25.7 +/- 0.8 mV (n = 67), reduces the apparent potassium selectivity of the cell membrane, from 0.55 +/- 0.07 (n = 9) to 0.09 +/- 0.01 (n = 26), and increases the apparent chloride selectivity from close to zero to 0.34 +/- 0.02 (n = 21). Potassium channel blocker barium (1 mmol/l) depolarizes the cell membrane by +15.2 +/- 1.1 mV (n = 13). In the presence of barium, reduction of extracellular osmolarity leads to a further depolarization by +14.0 +/- 1.4 mV (n = 12). Addition of chloride channel blocker anthracene-9-COOH (1 mmol/l) leads to a hyperpolarization of the cell membrane by -6.7 +/- 2.2 mV (n = 11). In the presence of anthracene-9-COOH, reduction of the extracellular osmolarity leads to a depolarization by +22.4 +/- 1.7 mV (n = 11).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Effect of extracellular adenosine triphosphate on electrical properties of subconfluent Madin-Darby canine kidney cells.

1. The present study has been performed to test for an influence of extracellular ATP on the potential differences across the cell membrane (PD) in subconfluent MDCK cells utilizing conventional microelectrodes. 2. In the absence of ATP, the mean measured PD was -47.5 +/- 0.3 mV (+/- S.E.M., n = 320). Application of 10 mumol/l ATP leads to rapid (less than 2 s) hyperpolarization of the cell membrane by -18.5 +/- 0.4 mV (n = 221), reduction of input resistance by 14 +/- 1 M omega (n = 106) and increase of the sensitivity of PD to alterations of extracellular potassium. 3. The concentration needed for half-maximal effect (K1/2) of ATP is approximately 0.5 mumol/l. ATP-gamma-S (K1/2 approximately 0.4 mumol/l) aand ADP (K1/2 approximately 0.9 similarly effective, whereas up to 1 mmol/l AMP or adenosine does not significantly alter PD. Application of 10 mumol/l theophylline, 1 mumol/l phentolamine and 10 mumol/l indomethacin does not blunt the hyperpolarizing effect of ATP. 4. The ATP-induced hyperpolarization is completely abolished in the presence of 1 mmol/l quinidine but only incompletely by 0.1 mmol/l quinidine or 1 mmol/l barium. In calcium-free extracellular fluid (1 mmol/l EDTA added) PD is 18.5 +/- 1.7 mV (n = 18). With reduced extracellular calcium, the hyperpolarizing effect of ATP is blunted (-12.3 +/- 1.6 mV, n = 18) and only transient. 5. In conclusion, ATP hyperpolarizes MDCK cells by increasing the potassium conductance. The activation of potassium channels requires calcium.

Adenosine Diphosphate↗
PubMed1989

Activation of potassium channels in renal epithelioid cells (MDCK) by extracellular ATP.

Extracellular ATP has been shown to stimulate transepithelial chloride transport in confluent Madin-Darby canine kidney (MDCK) cell layers and to enhance potassium conductance in subconfluent MDCK cells. The present study has been performed to test for the effect of extracellular ATP on channel activity in patches from subconfluent MDCK cells. Within 8 s, addition of extracellular ATP (10 mumol/l) leads to a sustained, but fully reversible, appearance of potassium-selective channels in cell-attached patches [increase of open probability from 0.03 +/- 0.02 (n = 10) to 0.50 +/- 0.07 (n = 6)]. With the use of pipettes filled with 145 mmol/l KCl, inwardly rectifying property of the channels is disclosed with a single-channel conductance of 65.7 +/- 3.1 pS (n = 9) at zero potential difference between pipette and bath and with a reversal potential of 75.4 +/- 2.0 mV (n = 5; pipette negative vs. reference in the bath). The open probability of the channels is not significantly modified by altering pipette potential from -50 mV, pipette positive, to 50 mV, pipette negative. At extracellular calcium activities of less than 10 nmol/l, ATP leads to a transient activation of channels. In conclusion, extracellular ATP activates inwardly rectifying potassium channels in the cell membrane of subconfluent MDCK cells. A sustained activation of the channels requires the presence of extracellular calcium and is probably mediated by increases in intracellular calcium.

Adenosine Triphosphate↗
PubMed1988

Enhancement of intracellular calcium concentration by extracellular ATP and UTP in Madin Darby Canine Kidney cells.

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.

Adenosine Triphosphate↗
PubMed1988

Effects of extracellular nucleotides on electrical properties of subconfluent Madin Darby canine kidney cells.

ATP and ADP but not AMP lead to sustained hyperpolarization of Madin Darby canine kidney (MDCK) cells. The present study has been performed to test for an influence of other nucleotides on the potential difference across the cell membrane (PD) in subconfluent MDCK cells. PD has been continuously monitored with conventional microelectrodes during rapid exchange of extracellular fluid. Application of 1 mumol/1 UTP leads to a rapid (less than 2 s) hyperpolarization of the cell membrane by -17.0 +/- 0.4 mV (from -50.1 +/- 0.6 mV), a reduction of cell membrane resistance and an increase of the sensitivity of PD to alterations of extracellular potassium. The concentration needed for half maximal effect of UTP is approximately equal to 0.2 mumol/1. ITP is similarly effective, whereas UDP, GTP and GDP are less effective. Up to 1 mmol/1 UMP, GMP, TTP or CTP do not significantly alter PD. In calcium-free extracellular fluid the hyperpolarizing effect of UTP is blunted (-11.6 +/- 2.3 mV) and only transient. In conclusion, UTP similar to purine triphosphates hyperpolarizes MDCK cells by increasing the potassium conductance. The activation of potassium channels requires calcium, which is apparently recruited from both intra- and extracellular sources.

Animals↗
PubMed1988

Pertussis toxin-dependent and -independent hormonal effects on cultured renal epithelioid cells.

The present study has been performed to test for the involvement of pertussis toxin-sensitive GTP-binding proteins (G-proteins) in the cellular transduction of hormone-induced activation of potassium channels. In Madin Darby canine kidney (MDCK) cells, a permanent cell line from dog kidney, epinephrine, acetylcholine, bradykinin, serotonin and ATP hyperpolarize the cell membrane by activation of potassium channels. In cells pretreated with pertussis toxin the hyperpolarizations elicited by either acetylcholine or serotonin are completely abolished; that following epinephrine is blunted and only transient. The hyperpolarizing effects of ATP or bradykinin are not affected by pertussis toxin. Thus, in MDCK cells both pertussis toxin-dependent and -independent mechanisms operate in parallel to enhance the potassium conductance of the cell membrane.

Acetylcholine↗
PubMed1988

Effect of acetylcholine on electrical properties of subconfluent Madin Darby canine kidney cells.

To elucidate the effects of acetylcholine on the electrical properties of incompletely confluent Madin Darby canine kidney (MDCK) cells continuous measurements of the potential difference across the cell membrane (PD) were made with conventional microelectrodes during rapid changes of extracellular fluid composition. During control conditions PD averages -48.9 +/- 1.0 mV (n = 51). 1 mumol/l acetylcholine leads to a sustained but reversible hyperpolarization of the cell membrane by -17.9 +/- 0.7 mV (n = 51). Half-maximal effect is observed at some 100 nmol/l. 1 mumol/l atropine does not significantly alter the potential difference across the cell membrane, but abolishes reversibly the hyperpolarizing effect of acetylcholine. Increase of extracellular potassium concentration from 5.4 mmol/l to 20 mmol/l depolarizes the cell membrane by +12.1 +/- 1.1 mV (n = 12) in the absence and by +25.7 +/- 0.9 mV (n = 12) in the presence of acetylcholine. Within 80 s removal of extracellular calcium leads to a depolarization of the cell membrane by +16.2 +/- 3.2 mV (n = 9). In the nominal absence of extracellular calcium acetylcholine leads to a transient hyperpolarization by -13.8 +/- 1.8 mV (n = 9), which can be elicited only once. In conclusion, acetylcholine hyperpolarizes the plasma membrane of MDCK cells by calcium-dependent enhancement of potassium conductance.

Acetylcholine↗
PubMed1988

Effects of serotonin on electrical properties of Madin-Darby canine kidney cells.

The present study has been performed to test for the influence of serotonin on the potential difference across the cell membrane (PD) of Madin-Darby canine kidney (MDCK)-cells. Under control conditions PD averages -48.6 +/- 0.6 mV (n = 98). Increasing extracellular potassium concentration from 5.4 to 10 and 20 mmol/l depolarizes the cell membrane by +6.3 +/- 0.6 mV (n = 6) and +14.1 +/- 1.0 mV (n = 12), respectively. The cell membrane is transiently hyperpolarized to -67.8 +/- 0.8 mV (n = 63) by 1 mumol/l serotonin. In the presence of serotonin, increasing extracellular potassium concentration from 5.4 to 20 mmol/l depolarizes the cell membrane by +26.4 +/- 1.0 mV (n = 11). 1 mmol/l barium depolarizes the cell membrane by +15.7 +/- 1.3 mV (n = 17) and abolishes the effect of step increases of extracellular potassium concentration from 5.4 to 10 mmol/l. In the presence of barium, serotonin leads to a transient hyperpolarization by -26.3 +/- 1.0 mV (n = 16). During this transient hyperpolarization, the cell membrane is sensitive to extracellular potassium concentration despite the continued presence of barium. 10 mumol/l methysergide hyperpolarize the cell membrane by -7.2 +/- 2.0 mV (n = 6). In the presence of 10 mumol/l methysergide, the effect of serotonin is virtually abolished (+0.4 +/- 0.9 mV, n = 6). 1 mumol/l ketanserin, a 5-HT2 receptor blocking agent, ICS 205-930, a 5-HT3 receptor blocking agent, and phentolamine, an unspecific alpha-receptor blocking agent, do not significantly modify the effect of serotonin. In the nominal absence of extracellular calcium, the effect of serotonin is markedly reduced.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Antagonists↗
PubMed1988

Inward rectifier K channels in renal epithelioid cells (MDCK) activated by serotonin.

The present study has been performed to test for the effect of intracellular calcium and of serotonin on the channel activity in patches from subconfluent MDCK-cells. In inside-out patches, inwardly rectifying potassium-selective channels are observed with open probabilities of 0.01 +/- 0.01, 0.24 +/- 0.03 and 0.39 +/- 0.07, at 100 nmol/liter, 1 mumol/liter or 10 mumol/liter calcium activity, respectively. The single-channel slope conductance is 34 +/- 2 pS, if the potential difference across the patch (Vp) is zero, and approaches 59 +/- 1 pS, if Vp is -50 mV, cell negative. In the cell-attached mode, little channel activity is observed prior to application of serotonin (open probability = 0.03 +/- 0.03). If 1 mumol/liter serotonin is added to the bath perfusate, the open probability increases rapidly to a peak value of 0.34 +/- 0.04 within 8 sec. In continued presence of the hormone, the open probability declines to approach 0.06 +/- 0.02 within 30 sec. At zero potential difference between pipette and reference in the bath (i.e., the potential difference across the patch is equal to the potential difference across the cell membrane), the single-channel conductance is 59 +/- 4 pS. In conclusion, inwardly rectifying potassium channels have been identified in the cell membrane of subconfluent MDCK-cells, which are activated to a similar extent by increase of intracellular calcium activity to 1 mumol/liter and by extracellular application of 1 mumol/liter serotonin.

Animals↗
PubMed1987

Effects of bradykinin on electrical properties of Madin-Darby canine kidney epithelioid cells.

In the present study we have investigated the influence of bradykinin on the potential difference across the cell membrane (PD) of Madin Darby Canine Kidney (MDCK)-cells. In the absence of bradykinin PD averages -52.6 +/- 0.9 mV (n = 52). Increasing extracellular potassium concentration from 5.4 to 10 and 20 mmol/l depolarizes the cell membrane by +5.2 +/- 0.3 mV (n = 8) and +14.9 +/- 1.0 mV (n = 9), respectively. The application of 0.1 mumol/l bradykinin leads to a transient hyperpolarization of the cell membrane to -70.3 +/- 0.6 mV (n = 30). During this transient hyperpolarization increasing extracellular potassium concentration from 5.4 to 10 and 20 mmol/l depolarizes the cell membrane by +10.4 +/- 0.7 mV (n = 10) and +29.2 +/- 0.8 mV (n = 8) respectively. Application of fragments of bradykinin (0.1 mumol/l) are without significant effect on the potential difference across the cell membrane. 1 mmol/l barium depolarizes the cell membrane by +15.8 +/- 1.2 mV (n = 9) and abolishes the effect of step increase of extracellular potassium concentration from 5.4 to 10 mmol/l. In the presence of barium, bradykinin leads to a transient hyperpolarization by -24.7 +/- 1.3 mV (n = 7). During this transient hyperpolarization, the cell membrane is sensitive to extracellular potassium concentration despite the continued presence of barium. In the nominal absence of extracellular calcium, bradykinin leads to a transient hyperpolarization, which can be elicited only once. The transient hyperpolarization is not affected by the presence of verapamil or indomethacin. In conclusion, bradykinin hyperpolarizes MDCK-cells by increasing the apparent potassium conductance.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Epinephrine activates outward rectifying K channel in Madin-Darby canine kidney cells.

Patch-clamp recordings were used to study the epinephrine dependent activation of ion channels in the cell membrane of cultured subconfluent renal epithelial (MDCK) cells. The patch-current was dominated by two populations of K channels. The spontaneously active population of K channels shows an inward rectifying behavior. Addition of epinephrine to the cell exterior, after the patch-pipette had been sealed to the cell membrane, increased the open probability of the inward rectifying K channel and shifted the membrane potential in the hyperpolarizing direction. The epinephrine induced hyperpolarization occurs in the range of seconds and is caused by activation of outward-rectifying K channels. The outward-rectifying K channel could not be observed under control conditions. Epinephrine activated channels always appeared in clusters of four to nine channels. Both populations of K channels are modulated in their open probability by cytoplasmic free calcium and voltage.

Animals↗
PubMed1987

Electrical properties of Ehrlich ascites tumor cells.

The cell membrane potential (PD) of Ehrlich ascites tumor cells was measured continuously at 37 degrees C with conventional microelectrodes during rapid alterations of extracellular fluid composition. At extracellular electrolyte composition mimicking the in vivo situation PD is -56.7 +/- 0.7 mV and the apparent membrane resistance is 62.2 +/- 2.2 M omega. Increasing extracellular potassium concentration from 5.4 to 20.0 mmol/l depolarizes the cell membrane by +18.4 +/- 0.5 mV. Thus, the transference number for potassium (tk, apparent slope potassium conductance over slope membrane conductance) is 0.53 +/- 0.01. A significant correlation is observed between tk and PD: tk = -(0.014 +/- 0.001) [1/mV] X PD [mV] -(0.243 +/- 0.051). 0.7 mmol/l barium depolarizes the cell membrane by +28.2 +/- 0.7 mV, increases the apparent membrane resistance by a factor of 2.6 +/- 0.1 and abolishes the apparent potassium conductance. Reduction of extracellular sodium concentration from 141 to 21 mmol/l depolarizes the cell membrane by +3.1 +/- 1.3 mV. Similarly, 0.1 mmol/l amiloride depolarizes the cell membrane by +3.3 +/- 0.7 mV. Reduction of extracellular chloride concentration from 128 to 67 mmol/l hyperpolarizes the cell membrane by -2.5 +/- 0.2 mV. 1 mmol/l anthracene-9-COOH does not significantly alter PD. Temporary omission of glucose from the extracellular fluid has no appreciable effect on PD. In conclusion, PD of Ehrlich ascites tumor cells is in the range of other mammalian epithelial cells and is generated mainly by potassium diffusion, while the conductances to sodium and chloride appear to be small.

Animals↗