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H Gögelein

Publications and source records attributed to H Gögelein.

At least 19 recordsLinked to original sources
PubMed1992

A calcium and ATP sensitive nonselective cation channel in the antiluminal membrane of rat cerebral capillary endothelial cells.

The patch-clamp technique was applied to the antiluminal membrane of freshly isolated capillaries of rat brain (blood-brain barrier). With 1.3 mM Ca2+ in the bath, excision of membrane patches evoked ion channels, which could not be observed in cell-attached mode. The channel was about equally permeable to Na+ and K+ ions, but not measurable permeable to Cl- and the divalent ions Ca2+ and Ba2+. The current-voltage curve was linear in the investigated voltage range (-80 mV to +80 mV), and the single-channel conductance was 31 +/- 2 pS (n = 22). The channel open probability was not dependent on the applied potential. Lowering of Ca2+ to 1 microM or below on the cytosolic side inactivated the channels, whereas addition of cytosolic ATP (1 mM) inhibited channel activity completely and reversibly. The channel was blocked by the inhibitor of nonselective cation channels in rat exocrine pancreas 3',5-dichlorodiphenylamine-2-carboxylic acid (DCDPC, 10 microM) and by the antiinflammatory drugs flufenamic acid (greater than 10 microM) and tenidap (100 microM), as well as by gadolinium (10 microM). Thus, these nonselective cation channels have many properties in common with similar channels observed in fluid secreting epithelia. The channel could be involved in the transport of K+ ions from brain to blood side.

Adenosine Triphosphate↗
PubMed1992

Activation of nonselective cation channels in the basolateral membrane of rat distal colon crypt cells by prostaglandin E2.

Ion channels in the basolateral membrane of colonic crypts were investigated with the patch-clamp technique during stimulation of secretion. Intact crypts were isolated from rat distal colon and the cell potential was recorded by addition of nystatin to the pipette solution. The cell resting potential in the base of the crypt was -74 +/- 1 mV (n = 90). Addition of 100 microM carbachol to the bath resulted in a transient hyperpolarization by 9 mV, which was probably due to the opening of basolateral K+ channels. In contrast, application of prostaglandin E2 (PGE2, 1 nM-1 microM) caused a dose-dependent depolarization in the base of the crypt. With 1 microM PGE2 cells depolarized from -74 +/- 1 to -27 +/- 2 mV (n = 26). Cell potential recordings in the midcrypt showed only a slight and transient depolarization after application of PGE2, whereas cells close to the surface of the crypt had no response. In the base of the crypt the PGE2-induced depolarization could be completely inhibited by addition of 50 microM flufenamic acid, a known blocker of nonselective cation channels. After substitution of all monovalent cations by N-methyl-D-glucamine in the bath, PGE2 had no significant effect on the cell potential. Cell-attached experiments with no nystatin in the patch pipette revealed the activation of ion channels in the basolateral membrane after application of PGE2. After excision of the membrane patch, these channels could be identified as nonselective cation channels. Experiments involving substitution of the bath solution showed that the channel is impermeable for Cl- and scarcely permeable for Ca2+ ions. The permeability sequence for monovalent cations, as calculated from reversal potentials, is NH4+ greater than Na+ = K+ greater than Rb+ = Li+ much greater than TRIS+ = NMDG+. Single channels are completely inhibited by flufenamic acid (50 microM), mefenamic acid (200 microM), as well as by 3',5-dichlorodiphenylamine-2-carboxylate. In conclusion, PGE2 activates nonselective cation channels in the basolateral membrane of cells in the base of colonic crypts. It is suggested that this mechanism initiates the secretion of K+ ions. Na+ influx through the nonselective cation channel will stimulate the Na+/K+ pump and active uptake of K+ at the basolateral side. K+ can leave the cell at the luminal side through K(+)-selective channels.

Animals↗
PubMed1992

Small and maxi K+ channels in the basolateral membrane of isolated crypts from rat distal colon: single-channel and slow whole-cell recordings.

The patch-clamp technique was used to characterize K+ channel activity in the basolateral membrane of isolated crypts from rat distal colon. In cell-attached patches with KCl in the pipette, channels with conductances ranging from 6 pS to 80 pS appeared. With NaCl in the pipette and KCl in the bath in excised inside-out membrane patches a small-conductance channel with a mean conductance of 12 +/- 6 pS (n = 18) was observed. The channel has been identified as K+ channel by its selectivity for K+ over Na+ and by its sensitivity to conventional K+ channel blockers, Ba2+ and tetraethylammonium (TEA+). Changes of cytosolic pH did not attenuate channel activity. Activity of the 12-pS channel was increased by membrane depolarization and elevated cytosolic Ca2+ concentration. In addition, a maxi K+ channel with a mean conductance of 187 +/- 15 pS (n = 4) in symmetrical KCl solutions was only occasionally recorded. The maxi K+ channel could be blocked by Ba2+ (5 mmol/l) on the cytosolic side. Using the slow-whole cell recording technique under control conditions, a cell membrane potential of -70 +/- 10 mV (n = 18) was measured. By application of various K+ channel blockers such as glibenclamide, charybdotoxin, apamin, risotilide, Ba2+ and TEA+ in the bath, only Ba2+ and TEA+ depolarized the cell membrane. The present data suggest that the small K+ channel (12 pS) is involved in the maintenance of the cell membrane resting potential.

Animals↗
PubMed1992

Stretch-activated non-selective cation channels in the antiluminal membrane of porcine cerebral capillaries.

1. Single stretch-activated (SA) channels have been studied in isolated brain capillary endothelial cells as well as in the antiluminal membrane of intact porcine cerebral capillaries using the patch-clamp recording technique. 2. The SA channels were found to be cation selective and permeable to Na+, K+, Ba2+ and Ca2+. 3. With monovalent cations in the patch pipette, the channels showed inward rectification in cell-attached patches with a single-channel conductance of 37 pS at negative and 24 pS at positive clamp potentials. 4. With either 70 mM-Ca2+ or Ba2+ in the patch pipette, the current-voltage relation was linear with slope conductances of 16 and 19 pS, respectively. 5. Mean channel open probability increased with increasing pressure and with depolarizing clamp potentials. 6. Cell swelling induced by hypotonic shock activated the SA channels in cell-attached experiments. 7. The SA channel may be involved in cell volume or blood flow regulation. The contribution of these channels to the regulation of cerebrospinal salt and water content, especially in brain oedema, is discussed.

Animals↗
PubMed1991

Angiotensin II, vasopressin and GTP[gamma-S] inhibit inward-rectifying K+ channels in porcine cerebral capillary endothelial cells.

Cerebral capillaries from porcine brain were isolated, and endothelial cells were grown in primary culture. The whole-cell tight seal patch-clamp method was applied to freshly isolated single endothelial cells, and cells which were held in culture up to one week. With high K+ solution in the patch pipette and in the bath we observed inward-rectifying K+ currents, showing a time-dependent decay in part of the experiments. Ba2+ (1-10 mM) in the bath blocked this current, whereas outside tetraethylammonium (10 mM) decreased the peak current but increased the steady-state current. Addition of 1 microM of angiotensin II or of arginine-vasopressin to the extracellular side caused a time-dependent inhibition of the inward-rectifying K+ current in part of the experiments. Addition of 100 microM GTP[gamma-S] to the patch pipette blocked the K+ inward rectifier. In cell-attached membrane patches two types of single inward-rectifying K+ channels were observed, with single channel conductances of 7 and 35 pS. Cell-attached patches were also obtained at the antiluminal membrane of intact isolated cerebral capillaries. Only one type of K+ channel with g = 30 pS was recorded. In conclusion, inwardly rectifying K+ channels, which can be inhibited by extracellular angiotensin II and arginine-vasopressin, are present in cerebral capillary endothelial cells. The inhibition of this K+ conductance by GTP[gamma-S] indicates that G-proteins are involved in channel regulation. It is suggested that angiotensin II and vasopressin regulate K+ transport across the blood-brain barrier, mediating their effects via G-proteins.

Angiotensin II↗
PubMed1991

Sodium-alanine cotransport in renal proximal tubule cells investigated by whole-cell current recording.

Sodium-alanine cotransport was investigated in single isolated proximal tubule cells from rabbit kidney with the whole-cell current recording technique. Addition of L-alanine at the extracellular side induced an inward-directed sodium current and a cell depolarization. The sodium-alanine cotransport current was stereospecific and sodium dependent. Competition experiments suggested a common cotransport system for L-alanine and L-phenylalanine. Sodium-alanine cotransport current followed simple Michaelis-Menten kinetics, with an apparent Km of 6.6 mM alanine and 11.6 mM sodium and a maximal cotransport current of 0.98 pA/pF at -60 mV clamp potential. Hill plots of cotransport current suggested a potential-independent coupling ratio of one sodium and one alanine. The apparent Km for sodium and the maximal cotransport current were potential dependent, whereas the apparent Km for L-alanine was not affected by transmembrane potential. The increase in Km for alanine with decreasing inward-directed sodium gradients suggested a simultaneous transport mechanism. These results are consistent with a cotransport model with potential-dependent binding or unbinding of sodium (high-field access channel) and a potential-dependent translocation step.

Alanine↗
PubMed1990

Quinine inhibits chloride and nonselective cation channels in isolated rat distal colon cells.

Isolated cells from rat distal colon were investigated with the patch-clamp technique. In cell-attached and cell-excised patches (inside-out) single chloride channels with outward-rectifying properties were observed. In excised patches the single-channel conductance g was 47 +/- 5 pS at positive and 22 +/- 2 pS at negative clamp potentials (n = 6). The Cl- channel blocker 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB, 10 microM) induced fast closing events, whereas 10 microM of 3',5-dichlorodiphenylamine-2-carboxylic acid (DCDPC) had no effect when applied to the cytosolic side. Quinine in the bath inhibited the Cl- channel by reducing its single-channel amplitude and increased open channel noise. With 0.1 mM the current amplitude decreased by 54% and with 1 mM quinine by 67%. Ca2(+)-dependent nonselective cation channels where observed after excision of the membrane patch. This channel was completely and reversibly inhibited by 100 microM DCDPC. Application of 1 mM quinine to the bath induced flickering and reduced the open-state probability from 0.94 to 0.44. In summary, besides its well established effects on K+ channels, quinine also inhibits nonselective cation channels and chloride channels by inducing fast closing events.

Animals↗
PubMed1990

Flufenamic acid, mefenamic acid and niflumic acid inhibit single nonselective cation channels in the rat exocrine pancreas.

The non-steroidal anti-inflammatory drugs, flufenamic acid, mefenamic acid and niflumic acid, block Ca2(+)-activated non-selective cation channels in inside-out patches from the basolateral membrane of rat exocrine pancreatic cells. Half-maximal inhibition was about 10 microM for flufenamic acid and mefenamic acid, whereas niflumic acid was less potent (IC50 about 50 microM). Indomethacin, aspirin, diltiazem and ibuprofen (100 microM) had not effect. It is concluded that the inhibitory effect of flufenamate, mefenamate and niflumate is dependent on the specific structure, consisting of two phenyl rings linked by an amino bridge.

Animals↗
PubMed1990

Voltage-dependent InsP3-insensitive calcium channels in membranes of pancreatic endoplasmic reticulum vesicles.

Stimulus-secretion coupling in exocrine glands involves Ca2+ release from intracellular stores. In endoplasmic reticulum vesicle preparations from rat exocrine pancreas, an inositol 1,4,5-trisphosphate(InsP3)-sensitive, as well as an InsP3-insensitive, Ca2+ pool has been characterized. But Ca2+ channels in the endoplasmic reticulum of rat exocrine pancreas have not been demonstrated at the level of single-channel current. We have now used the patch-clamp technique on endoplasmic reticulum vesicles fused by means of the dehydration-rehydration method. In excised patches, single Ba2(+)- and Ca2(+)-selective channels were recorded. The channel activity was markedly voltage-dependent. Caffeine increased channel open-state probability, whereas ruthenium red and Cd2+ blocked single-channel currents. Ryanodine, nifedipine and heparin had no effect on channel activity. The channel activity was not dependent on the free Ca2+ concentration, the presence of InsP3, or pH. We conclude that this calcium channel mediates Ca2+ release from an intracellular store through an InsP3-insensitive mechanism.

Animals↗
PubMed1989

The nonselective cation channel in the basolateral membrane of rat exocrine pancreas. Inhibition by 3',5-dichlorodiphenylamine-2-carboxylic acid (DCDPC) and activation by stilbene disulfonates.

Nonselective Ca2+-sensitive cation channels in the basolateral membrane of isolated cells of the rat exocrine pancreas were investigated with the patch clamp technique. With 1.3 mmol/l Ca2+ on the cytosolic side, the mean open-state probability Po of one channel was about 0.5. In inside-out oriented cell-excised membrane patches the substances diphenylamine-2-carboxylic acid (DPC), 5-nitro-2-(3-phenelpropylamino)-benzoic acid (NPPB) and 3',5-dichlorodiphenylamine-2-carboxylic acid (DCDPC) were applied to the cytosolic side. These compounds inhibited the nonselective cation channels by increasing the mean channel closed time (slow block). 100 mumol/l of NPPB or DPC decreased Po from 0.5 (control conditions) to 0.2 and 0.04, respectively, whereas 100 mumol/l of DCDPC blocked the channel completely. All effects were reversible. 1 mmol/l quinine also reduced Po, but in contrast to the above mentioned substances, it induced fast flickering. Ba2+ (70 mmol/l) and tetraethylammonium (TEA+; 20 mmol/l) had no effects. We investigated also the stilbene disulfonates 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS), 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) and 4,4'-dinitro-2,2'-stilbenedisulfonate (DNDS). 10 mumol/l SITS applied to the cytosolic side increased Po from 0.5 to 0.7 and with 100 mumol/l SITS the channels remained nearly permanently in its open state (Po approximately equal to 1). A similar activation of the channels was also observed with DIDS and DNDS. These effects were poorly reversible. The stilbene disulfonates acted by increasing the channel mean open time. When the channel was inactivated by decreasing bath Ca2+ concentration to 0.1 mumol/l, addition of 100 mumol/l of SITS had no effect. Similarly, reducing bath Ca2+ concentration from 1.3 mmol/l in presence of 100 mumol/l SITS (channels are maximally activated) to 0.1 mumol/l, inactivated the channels completely. These results demonstrate, that SITS can only activate the channels in the presence of Ca2+. SITS had no effects, when applied to the extracellular side in out-side out patches.(ABSTRACT TRUNCATED AT 400 WORDS)

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1989

Versatile supplement device with remote control for the control of patch clamp experiments.

A versatile device for a patch-clamp amplifier is described. This device contains: (i) an acoustic indicator to monitor the input resistance of the patch pipette, which is used in search-mode to indicate the formation of seals; (ii) two pulse generators; and (iii) a staircase generator to produce various pulse and voltage step programs; (iv) a low-pass filter which is used to filter the output of the patch clamp amplifier; and (v) a remote control which is used to control the entire patch clamp experiment. This remote control is used to switch between search-, current clamp-, and voltage clamp-mode, to activate the respective stimulus potential programs, and to control the tape recorder. This electronic device can be easily connected to patch clamp amplifiers.

Animals↗
PubMed1989

Single chloride channels in endosomal vesicle preparations from rat kidney cortex.

Endocytotic vesicles from rat kidney cortex, isolated by differential centrifugation and enriched on a Percoll gradient, contain both an electrogenic H+ translocation system and a conductive chloride pathway. Using the dehydration/rehydration method, we fused vesicles of enriched endosomal vesicle preparations and thereby made them accessible to the patch-clamp technique. In the fused vesicles, we observed Cl- channels with a single-channel conductance of 73 +/- 2 pS in symmetrical 140 mM KCl solution (n = 25). The current-voltage relationship was linear in the range of -60 to +80 mV, but channel kinetic properties depended on the clamp potential. At positive potentials, two sublevels of conductance were discernible and the mean open time of the channel was 10-15 msec. At negative voltages, only one substate could be resolved and the mean open time decreased to 2-6 msec. Clamp voltages more negative than -50 mV caused reversible channel inactivation. The channel was selective for anions over cations. Ion substitution experiments revealed an anion permeability sequence of Cl- = Br- = I- greater than SO4(2-) approximately F-. Gluconate, methanesulfonate and cyclamate were impermeable. The anion channel blockers 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS, 1.0 mM) and 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB, 0.1 mM) totally inhibited channel activity. Comparisons with data obtained from radiolabeled Cl(-)-flux measurements and studies on the H+ pump activity in endocytotic vesicle suspensions suggest that the channel described here is involved in maintenance of electroneutrality during ATP-driven H+ uptake into the endosomes.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1988

Blocking kinetics of Cl- channels in colonic carcinoma cells (HT29) as revealed by 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB).

The blocking effect of 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) was investigated on single Cl- channels of the cultured human colon carcinoma cells, HT29. In the absence of NPPB, the open-time histogram yielded two time constants, with 0.9 ms and 33 ms, whereas the closed-time distribution could be fitted by a single exponential with a time constant of 0.7 ms. Addition of NPPB in the range 1-50 microM induced brief closing events of the single-channel current. This resulted in a decrease of the long open-time constant to 2.1 ms and in an increase of the closed-time constant to 1.8 ms at 50 microM NPPB concentration. The short open-time constant did not change at low blocker concentration (1 microM), but could no longer be resolved at higher concentrations. The open-state probability decreased from 0.9 (control conditions) to 0.5 at 50 microM NPPB. The Hill plot yielded a Hill coefficient of about 0.7, compatible with one NPPB molecule inhibiting one channel molecule. The kinetics of channel gating are described by a sequential model with one closed and two open states. Since in the presence of NPPB no additional time constant appeared in the time histograms, we assumed the same kinetic scheme as under control conditions, and hypothesize that NPPB has an influence on rate constants.

Cell Line↗
PubMed1988

Anion channels in giant liposomes made of endoplasmic reticulum vesicles from rat exocrine pancreas.

Using the method of dehydration and rehydration, rough endoplasmic reticulum (RER) vesicles, isolated by differential centrifugation, can be enlarged to giant liposomes with diameters ranging from 5 to 200 micron. Patch-clamp studies on these giant RER liposomes revealed the existence of a channel with a mean conductance of 260 +/- 7 pS (n = 23; 140 mmol/liter KCl on both sides). The channel is about four times more permeable for Cl- than for K+. Its activity is strongly voltage regulated. At low potentials (+/- 20 mV) the channel is predominantly in its open state with an open probability near 1.0, whereas it closes permanently at high positive and negative voltages (+/- 70 mV). The channel activity is not influenced by changing the free Ca2+ concentration from 1 mmol/liter to less than 10(-9) mol/liter on either side, and is also not affected by typical Cl- -channel blockers like diphenylamine-2-carboxylate (DPC, 1 mmol/liter) or 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS, 1 mmol/liter). Another chloride channel with a single-channel conductance of 79 +/- 6 pS (n = 4) was less frequently observed. In the potential range of -80 to +40 mV this channel displayed no voltage-dependent gating. We assume that these anion channels are involved in the maintenance of electroneutrality during Ca2+ uptake in the RER.

Animals↗
PubMed1988

Stimulation of NaCl secretion in the rectal gland of the dogfish Squalus acanthias.

1. The rectal gland of the dogfish (Squalus acanthias) secretes NaCl when stimulated by a hormone related to vasointestinal peptide. 2. Patch clamp and microelectrode techniques are used to examine the changes in membrane conductances occurring with hormonal stimulation. 3. The conductance of the "resting" cell is dominated by basolateral K+ channels. 4. Hormonal stimulation "opens" apical Cl- channels. 5. This opening of apical chloride channels appears to be mediated by cAMP-dependent phosphorylation of pre-existing closed channels.

Animals↗
PubMed1987

Properties of single K+ channels in the basolateral membrane of rabbit proximal straight tubules.

The basolateral membrane of rabbit straight proximal tubules, which were cannulated and perfused on one side, was investigated with the patch clamp technique. Properties of inward and outward directed single K+ channel currents were studied in cell-attached and inside-out oriented cell-excised membrane patches. In cell-attached patches with NaCl Ringer solution both in pipette and bath, outward K+ currents could be detected after depolarization of the membrane patch by about 20-30 mV. The current-voltage (i/V) relationship could be fitted by the Goldman-Hodgkin-Katz (GHK) current equation, with the assumption that these channels were mainly permeable for K+ ions. A permeability coefficient PK of (0.17 +/- 0.04).10(-12) cm3/s was obtained, the single channel slope conductance at infinite positive potential g(V infinity) was 50 +/- 12 pS and the single channel conductance at the membrane resting potential g(Vbl) was 12 +/- 3 pS (n = 4). In cell-excised patches, with NaCl in the pipette and KCl in the bath, the data could also be fitted to the GHK equation and yielded PK = (0.1 +/- 0.01).10(-12) cm3/s, g(V infinity) = 40 +/- 4 pS and g(Vbl) = 7 +/- 1 pS (n = 8). In cell-attached patches with KCl in the pipette and NaCl in the bath, inward K+ channels occurred at clamp potentials less than or equal to 60 mV, whereas outward K+ channel current was detected at more positive voltages. The current-voltage curves showed slight inward rectification.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Potassium channels in the basolateral membrane of the rectal gland of the dogfish (Squalus acanthias).

Previous studies in isolated, in vitro perfused rectal gland tubules (RGT) have revealed that the basolateral membrane possesses a K+ conductive pathway. In the present study, we have utilized the patch clamp technique in RGT segments to characterize this pathway. The basolateral membrane was approached with patch pipettes at the open end of in vitro perfused segments. Recordings were obtained in cell-attached as well as in excised inside-out patches. In cell-attached patches with the pipette filled with a KCl solution (274 mmol/l) and the bath containing NaCl shark Ringer (275 mmol/l), inward K+ currents (from pipette into cell) with a mean slope conductance of 123 +/- 26 pS (n = 3) were observed. We were unable to generate outward K+ currents at high depolarizing (cell more positive) clamp voltages. This indicates inward rectification of this channel. To examine the rectification properties further, excised (inside out) patches were exposed to K+ concentration gradients, directed out of, as well as into the pipette. With NaCl in the pipette and KCl in the bath, K+ outward currents were observed. The current-voltage (IV) relation revealed Goldman-type rectification, with a mean single channel conductance of 185 +/- 28 pS (n = 7) at high positive voltages (linear range of the IV curve). The single-channel permeability coefficient for K+ was 0.26 +/- 0.04 X 10(-12) cm3/s (n = 7). In the reversed experiment (pipette KCl, bath NaCl), inward currents of similar kinetics and amplitude were obtained. The single channel conductance was 146 +/- 21 pS (n = 7) at high negative voltages (linear range of the IV curve).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗