Galanin activates an inwardly rectifying potassium conductance in mudpuppy atrial myocytes

Galanin- and bethanechol-activated K⁺ currents have been studied in mudpuppy atrial myocytes. The galanin and bethanechol K⁺ currents were time-dependent and inwardly rectifying. In GTPS, the galanin and bethanechol currents were reduced progressively as G-protein gated K⁺ channels became activated. GDPS inhibited agonist-induced outward currents. We conclude that galanin and bethanechol activate the same or a very similar inwardly rectifying K⁺ conductance and that activation of a G protein is required.     

Transient effects of norepinephrine on myocardial oxygen balance

Summary In conscious dogs with experimental atrioventricular block and with ventricles paced at constant rate the effects of norepinephrine (NE) and isoproterenol (ISO) on coronary flow, coronary resistance, and myocardial O₂-balance were investigated. Myocardial O₂-s balance, as estimated from continuous measurement of coronary venous O₂-s saturation, was used for the discrimination of coronary dilation induced either directly by vascular -adrenoreceptor stimulation or indirectly by increased myocardial metabolism.Following bolus injection of NE (0.3 g/kg) or ISO (0.1 g/kg) into the pulmonary artery, coronary venous O₂-s saturation increased from a control of 25±2% O₂-s saturation (mean±S.D.) transiently to 51±5 and 62±5% O₂-s saturation respectively. After ₁-adrenoreceptor blockade these increases were reduced to 33±4 and 41±3% O₂-s saturation, respectively. The remaining increase after NE was abolished when atropine was given in addition to ₁-b blockade. After ₁+2-adrenoreceptor blockade neither NE nor ISO injection had an effect on coronary venous O₂ saturation. After ₁-b blockade was superimposed on ganglionic blockade NE injection led to a decrease in coronary venous O₂-s saturation indicating a ratent -a activity of NE.NE seems to act directly via ₁-a adrenoreceptors, since no differences were observed in the time courses of changes in coronary venous O₂-s saturation after left atrial injection of NE when compared to adenosine.It is concluded that circulating NE like ISO is able to improve myocardial oxygen balance by a direct vasodilating effect on canine coronary vessels mediated by vascular ₁-adrenoreceptors.     

An electrophysiological investigation of the characteristics and function of GABAA receptors on bovine adrenomedullary chromaffin cells

The characteristics and function of -aminobutyric acid_A (GABA_A) receptors expressed on bovine chromaffin cells in culture have been investigated using patch-clamp techniques. In voltage-clamped whole-cells, locally applied GABA (100 M) evoked a transmembrane chloride current which demonstrated outward rectification. The amplitude of such currents was reversibly suppressed by the GABA_A receptor antagonists bicuculline, picrotoxin and RU5135, and enhanced by the general anaesthetic propanidid. Glycine (100 M) and baclofen (100 M) were ineffective as agonists. In support of a physiological role for GABA in the adrenal medulla, the co-existence of GABA_A and nicotinic acetylcholine (ACh) receptors was demonstrated on whole cells and outside-out membrane patches. Ionophoretically applied GABA reduced the amplitude of depolarization and action potential discharge occurring in response to locally applied ACh (100 M), but had no effect upon the underlying ACh-induced current. In addition, an excitatory action of GABA was demonstrated by recording action potential waveforms in cell-attached patches. The results are discussed in the context of a GABA-ergic regulation of catecholamine secretion.     

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 mol/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 mol/l methysergide hyperpolarize the cell membrane by –7.2±2.0 mV (n=6). In the presence of 10mol/l methysergide, the effect of serotonin is virtually abolished (+0.4±0.9 mV,n=6). 1 mol/l ketanserin, a 5-HT₂ receptor blocking agent, ICS 205-930, a 5-HT₃ receptor blocking agent, and phentolamine, an unspecific -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. In conclusion, serotonin hyperpolarizes MDCK-cells by increasing apparent potassium conductance. This effect is transmitted by 5-HT₁ receptors and depends on extracellular calcium.     

Desensitization of the bradykinin-induced rise in intracellular free calcium in cultured endothelial cells

We studied the cellular mechanism involved in the desensitization of cultured endothelial cells to bradykinin. Bradykinin (10 nmol/l) evoked a rise in the intracellular free calcium concentration ([Ca _i ²⁺ ]), measured with the fluorescent probe indo-1, from 137±30 (±SEM) to 623±101 nmol/l. Cells were desensitized to bradykinin by repetitive stimulation with the peptide over 10 min, after which they no longer responded to bradykinin. However, purinergic stimulation with ATP (10 mol/l) elicited the same increase in [Ca _i ²⁺ ] in endothelial cells desensitized to bradykinin as in cells never exposed to bradykinin. The initial peak of [Ca _i ²⁺ ] after stimulation with bradykinin or ATP was not affected by removal of extracellular calcium ions, indicating mobilization of Ca²⁺ from intracellular stores. Since GTP-binding proteins (G-proteins) are probably involved in the receptor-mediated stimulation of endothelial cells, we also tested the effects of sodium fluoride (NaF), a reported direct stimulator of G-proteins, on endothelial [Ca _i ²⁺ ]. NaF (5 mmol/l) increased [Ca _i ²⁺ ] to 412±88 nmol/l in control cells and was equally effective in cells desensitized to bradykinin. We conclude that the homologous desensitization to bradykinin does not occur at the level of intracellular signal transduction but at the level of membrane receptors.     

Skull radiograph measurements of normals and patients with basilar impression; use of Landzert's angle

Summary One hundred normal lateral skull radiographs were studied and those of ten patients with basilar impression attending Kenyatta Hospital, Nairobi. The mean shortest distance of the odontoid tip to McGregor's basal line was 1.2±2.28 mm below the basal line (range 6 mm below to 3 mm above basal line), in normals and 9±2.7 mm (6–14 mm) above basal line in patients. The mean basal angle was 113±7 (102–133) in normals and 122±6 (113–125) in patients. The mean nasion-basion-opisthion angle was 162±4 (154–169) in normals and 178±5 (173–185) in patients. The mean total length of clivus was 48±3.7 mm (43–56 mm) in normals and 44±6.6 (36–48 mm) in patients group. The mean median diameter of the foramen magnum was 39±5 mm (30–48 mm), atlas 21±3 mm (18–25 mm) axis 18±3 mm (14–23 mm), third cervical vertebra 16±2 mm (13–22 mm) in normals and in patients: 39±4 mm (36–45 mm), atlas 23±6 (l5–30 mm) axis 19±4 mm (16–25 mm), third cervical vertebra 16±3 (14–20). There was a significant difference in the position of the odontoid tip and the nasion-basion-opisthion angle between the normal and patient groups. All the other parameters measured in this work did not differ significantly between the two groups.

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Etude anatomo-radiologique de crânes normaux et de crânes pathologiques avec impression basilaire; utilisation de l'angle de Landzert

Résumé Cent crânes normaux ont été étudiés sur des radiographies de profil ainsi que dix crânes pathologiques présentant des impressions basilaires chez des patients traités à l'HÔpital Kenyatta de Nairobi. La plus courte distance moyenne entre le sommet de l'odontoÏde et la ligne basale de McGregor a été de 1,2±2,28 mm au-dessous de la ligne basale (extrÊmes étendues de 6 mm au-dessous à 3 mm au-dessus de la ligne basale), chez les sujets normaux et de 9±2,7 mm (6–14 mm) au-dessus de la ligne basale chez les sujets pathologiques. L'angle basai moyen était de 113±7 (102–133) chez les sujets normaux et 122±6 (113–125) chez les sujets pathologiques. L'angle moyen nasion-basion-opisthion était de 162±4 (154–169) chez les sujets normaux et 178±5 (173–185) chez les sujets pathologiques. La longueur moyenne totale du clivus était de 48±3,7 mm (43–56 mm) chez les sujets normaux et 44±6,6 (36–48 mm) chez les sujets pathologiques. Le diamètre moyen du foramen magnum était de 39±5 mm (30–48 mm), celui du foramen vertébral de l'atlas était de 21±3 mm (18 à 25 mm), celui de l'axis (18±3 mm (14–23 mm), celui de la troisième vertèbre cervicale: 16±3 mm (13–22 mm) chez les sujets normaux; chez les sujets pathologiques les chiffres étaient les suivants: foramen magnum 39±4 mm (39–45 mm), atlas 23±6 (15–30 mm), axis 19±4 mm (16–25 mm), troisième vertèbre cervicale 16±3 mm (14–20 mm). Il existe une différence significative dans la position du sommet de l'odontoÏde et la valeur de l'angle nasion-basion-opisthion entre les deux groupes. Aucun des autres paramètres mesurés dans ce travail ne présentait de différence significative entre les deux groupes.

     

β-Subunits co-determine the sensitivity of rat neuronal nicotinic receptors to antagonists

We have investigated the effect of 4 ganglionic cholinergic antagonists (hexamethonium, mecamylamine, pentolinium, trimetaphan) on rat 32 and 34 neuronal nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus oocytes. Current responses were elicited by fast application of acetylcholine on voltage-clamped oocytes (holding potential Vinh = -80mV). Concentration-inhibition curves were used to get estimates of IC₅₀, the antagonist concentration yielding 50% reduction of the peak current. The K_B's of the antagonists were calculated using estimates of the apparent K_D of acetylcholine. The order of affinity of the antagonists was similar for both receptor subtypes: mecamylamine pentolinium > hexamethonium > trimetaphan. However, 34 neuronal nAChRs were 9 to 22 times more sensitive to each of the 4 antagonists than 32 receptors. These results further underline the importance of the -subunit as co-determinant of the functional properties of neuronal nAChRs.     

Pertussis-toxin-sensitive inhibition by (-) baclofen of Ca signals in bovine chromaffin cells

Ca signals in bovine adrenal chromaffin cells were studied both in Fura-2/AM-loaded intact cells, and in voltage-clamped cells under whole-cell patch-clamp conditions. The effects of gamma-aminobutyric acid b subtype (GABA_b) receptor activation on K⁺-depolarization-induced signals and on voltage-activated Ca²⁺ currents were investigated. Both GABA (20 M) plus bicuculline (20 M) and (-)baclofen (20–100 M), effectively inhibited the Ca signal in intact cells. The effects caused by baclofen continued to develop during the time interval between two successive stimuli. The restoration of the Ca signal during washout of baclofen was also delayed and continued in some experiments for 10–20 min. The inhibitory effect of baclofen on the Ca signal was eliminated by pre-treatment of the cells with pertussis toxin (PTX, 1g/ml, for 4–6h at 37°C). Baclofen (50 M) inhibited Ca²⁺ current in whole-cell mode by at most 20%. The effect developed quickly and was reversible. Infusion into the cells of a non-hydrolyzable analogue of guanosine 5-triphosphate GTP S (100 M), led to complete inhibition of the Ca²⁺ conductance and of voltage-evoked intracellular Ca ([CA]_i) transients within 2 min. In paired cells intracellularly perfused with GTPS-free solution, the Ca²⁺ current amplitude decreased by only about 30% for 5–6 min. It is concluded that bovine chromaffin cells have functional GABA_b receptors the activation of which, mediated by a PTX-sensitive GTP-binding protein, inhibits the evoked increase in cytosolic free Ca²⁺. The small size of the effect on Ca²⁺ current in whole-cell mode as compared to that on the Ca signal in intact cells may be explained by washout of some regulatory element during cell dialysis, or by a relatively small contribution of the normal voltage-activated Ca²⁺ current to the Ca signal. Alternatively, it might indicate GABA_b effects on mechanisms other than Ca²⁺ channels.     

Isolation of the Bα and Bβ mating-type loci of Schizophyllum commune

The B mating type of the basidiomycete fungus, Schizophyllum commune is determined by two, tightly linked, multi-specificity (also called multi-allelic) loci: B and B. A plasmid library was used in DNA-mediated transformation to obtain transformants that displayed B-directed development. Plasmids that conferred B1 and B1 mating-type specificities were rescued from the transformants. Fragments of DNA from each plasmid hybridized to genomic DNA from the strain used to make the plasmid library; however, they did not hybridize, or hybridized only weakly, to genomic DNA from strains with mating-type specificities different from B1 or B1. The cloned fragments are presumed to correspond to active regions of each B mating-type locus.     

An electrophysiological study of angiotensin II regulation of Na-HCO3 cotransport and K conductance in renal proximal tubules

The effect of picomolar concentrations of angiotensin II (AII) was investigated in isolated perfused rabbit renal proximal tubules using conventional or pH-sensitive intracellular microelectrodes. Under control conditions cell membrane potential (V _b) and cell pH (pH_i) averaged –53.8±1.9 mV (mean±SEM,n=49) and 7.24±0.01 (n=10), respectively. AII (at 10^–11 mol/l), when applied from the bath (but not when applied from the lumen perfusate), produced the following effects: approximately 85% of the viable tubules responded with a small depolarization (+ 5.5±0.4 mV,n=43) which was accompanied in half of the pH_i measurements by a slow acidification (pH_i=–0.03±0.01,n=5). The remaining 15% responded with a small hyperpolarization (V_b=–3.1±0.4 mV,n=6). All changes were fully reversible and repeatable. Experiments with fast changes in bath HCO₃ or K concentrations, as well as measurements of the basolateral voltage divider fraction in response to transepithelial current flow, explain these observations as stimulation of a basolateral Na-HCO₃ cotransporter and of a basolateral K conductance. Both counteract in their effect onV _b, but can be individuated by blocker experiments with 4,4-diisothiocyanatostilbene-2,2-disulphonic acid (DIDS) and barium. Both the stimulation of Na-HCO₃ cotransport and the stimulation of the K conductance may result from down-regulation of the level of cyclic adenosine monophosphate in the cell.     

Comparison of in vitro effects of flunixin and tolfenamic acid on human leukocyte and platelet functions

A study was made to compare the effects of two nonsteroidal antiinflammatory drugs (NSAIDs), flunixin and tolfenamic acid, on the leukotriene B₄ (LTB₄) production and migration of human polymorphonuclear leukocytes (PMNs) as well as on platelet aggregation and thromboxane B₂ (TxB₂) production during blood clotting. Tolfenamic acid inhibited LTB₄ production in PMNs as well as FMLP- and LTB₄-induced PMN migration (IC₅₀ values 23 ± 3, 39 ± 11, and 68 ± 13 M, respectively), whereas flunixin inhibited these cell functions only with the highest concentration tested (100 M). On the other hand, flunixin was clearly a more potent inhibitor of TxB₂ production and adrenaline-induced platelet aggregation than tolfenamic acid, the IC₅₀ values in TxB₂ production being 0.28 ± 0.02 M and 2.6 ±0.3 M for flunixin and tolfenamic acid, respectively. We suggest that inhibition of PMN functions may be an additional mechanism in the antiinflammatory action of tolfenamic acid. At least in human PMNs and platelets, flunixin seems to be only an inhibitor of cyclooxygenase.     

Changes in extracellular K+ evoked by GABA,THIP and baclofen in the guinea-pig hippocampal slice

Summary Changes in [K⁺]₀ evoked by the inhibitory amino acid transmitter, GABA (-aminobutyric acid) and its agonists were recorded with ion-selective microelectrodes in the CA1 stratum pyramidale of guinea-pig hippocampal slices. Bath applications of GABA (0.1–10 mM) produced dose-dependent increases in [K⁺]₀ (EC₅₀ = 4 mM, R_max= 1.6 mM), with a peak and decline during exposure, followed by undershoot during recovery. In contrast the selective GABA_a agonist, THIP (4,5,6,7-tetrahydroisoxazolo-(5,4-c)-pyridin-3-ol) (0.01–1 mM) showed ten-fold greater potency and evoked only increases in [K⁺]₀ (EC₅₀ = 0.5 mM, R_max= 2 mM). Reduction of temperature from 34° to 22° C caused a more than two-fold augmentation of the K₀ accumulation evoked by GABA, but no change in that due to THIP. The GABA_A antagonist, BMI (bicuculline methiodide) (100 M) completely blocked responses to THIP and partially antagonized those to GABA. Responses to GABA were synergistically enhanced by pentobarbital (100 M). Only small, delayed and inconsistent changes could be evoked by relatively high concentrations of the GABA_B agonist, DL-baclofen (0.01–1 mM). The K⁺ changes evoked by GABA appear to be mediated by the activation of GABA_A receptors with low affinity and to be related to their depolarizing action. Although the response includes an electrogenic component which suggests the involvement of Na-dependent transmitter uptake/transport, the increase in k ₀ ⁺ probably reflects an outward counter/co-transport of K⁺ with Cl/HCO₃ anion shifts and/or activation of a voltage-dependent K⁺ conductance.     

Naloxone in treatment of circulatory shock resistant to conventional therapy

Summary The effect of naloxone (4.4–5.9 mg i.v.) was evaluated in 10 patients with circulatory shock (sepsis,n=7; intoxication,n=1; cardiogenic shock,n=2) not responding to full conventional therapy. In addition, we measured plasma ACTH and immunoreactive -endorphin before and 60 min after administration of naloxone and compared the results with hormone concentrations in 10 intensive care patients without shock. Only in two patient with septic shock a transient increase (duration 15 min and 60 min, respectively) of systolic blood pressure was observed, while naloxone was ineffective in the remaining eight patients. No adverse effects of naloxone were found. Plasma ACTH and immunoreactive -endorphin concentrations in patients with shock were not different from those in controls (ACTH, 79±28 vs 120±60 pg/ml; immunoreactive -endorphin, 952±262 vs 1,070±378 pg/ml).Our findings suggest that naloxone in a single dose of 4.4–5.9 mg i.v. does not improve the management of circulatory shock unresponsive to conventional treatment. -endorphin seems to play no major role in the hypotension of shock.Abbreviations ACTH Adrenocorticotrophic hormone - HD intermittent hemodialysis - HF heart rate - ir immunoreactive - RR_syst systolic blood pressure Supported by Landesamt für Forschung, NRW     

A mouse mutant strain highly resistant to methyl β-carboline-3-carboxylate-induced seizures

The convulsant properties of methyl -carboline-3-carboxylate (-CCM) were evaluated in the TaT-fm/Gnc ^Ta+/+Tfm strain carrying the tabby coat color (Ta) and/or the testicular feminization (Tfm) gene. When injected intraperitoneally within a 5–60 mg/kg dose range, -CCM-induced convulsions in less than 25% of the mice, thus providing evidence for a high resistance of this strain, as compared to classical strains of mice. However, this strain responds normally to the convulsant pentylenetetrazol (PTZ), suggesting a specific resistance to -CCM. Both the Ta gene and the TaTfm/Gnc genetic background were involved in the high resistance to -CCM. In addition, concentrations of neurosteroids and benzodiazepine binding, both modulating GABA_A receptor efficacy, have been measured in order to elucidate the biological mechanisms of drug resistance.     

High-voltage-activated calcium current in developing neurons is insensitive to nifedipine

We have analyzed the effect of nifedipine on the macroscopic high-threshold, voltage-activated (HVA) calcium current in four cell types: postnatal rat Purkinje and dorsal root ganglion (DRG) neurons, embryonic chick DRG neurons, and adult cat ventricular myocytes. As is consistent with previous reports, nifedipine reduced HVA current in myocytes in a voltage-sensitive manner. Analysis of nifedipine actions on neurons, however, was compromised by slow inactivation of the current at holding potentials between –80mV and –40 mV. The slow inactivation was voltage-dependent, irreversible after 5 min, and contributed to rundown of the current. At –40 mV, slow inactivation displayed two time constants: 12±8 s and 7±4 min. When slow inactivation was taken into account, we found no evidence for a nifedipine-sensitive component of the HVA current in these neurons. Consistent with previous studies, DRG neurons were reduced irreversibly by -conotoxin, whereas cardiac and Purkinje cells were unaffected. Our biophysical and pharmacological results are consistent with two types of neuronal HVA currents (N type and P type) in developing neurons that are distinct from cardiac HVA currents (L type).     

A combined study of sodium current and T-type calcium current in isolated cardiac cells

Sodium currents (I _Na) and T-type calcium currents (I _Ca,T) of isolated guinea-pig ventricular myocytes were recorded using the whole-cell voltage-clamp technique. Separation of the two currents was obtained by using the difference current method in the presence and absence of 2 mM extracellular Na (Na_o). Time to peak and the time constant of inactivation of. I_Na were about 5 times faster than that of I _Ca,T (test potential –30 mV). and I _Ca,T had an activation range positive to –50 mV, were inactivated at –50 mV, and their current/voltage relationships peaked at –22.3±1.8 mV (n=18) and –29.3±0.5 mV (n=18) respectively, with a reversal potential of +40.3±4 mV (n=18) and +30±10 mV (n=18), respectively [2 mM Na_o; 5.4 mM extracellular Ca (Ca_o)]. I _Na was blocked by 30 M tetrodotoxin (TTX), 500 M lidocaine, partly inhibited by 1 mM amiloride, but not affected by 100 M nickel (Ni). I _Ca,T was neither affected by 30 M TTX nor 500 M lidocaine, but blocked by 100 M Ni, 1 mM amiloride, 10 M R 56865 and use-dependently reduced by 5 M flunarizine. Adenosine (500 M) affected neither I _Na nor I _Ca,T, whereas 1 M isoprenaline did not affect I _Ca,T, but slightly increased I _Na. Our results demonstrate that the characteristics of I _Ca,T are not affected by the concomitant activation of I _Na, and vice versa. We conclude that I _Ca,T are not Ca currents through Na channels.     

The luminal K+ channel of the thick ascending limb of Henle's loop

In vitro perfused rat thick ascending limbs of Henle's loop (TAL) were used (n=260) to analyse the conductance properties of the luminal membrane applying the patch-clamp technique. Medullary (mTAL) and cortical (cTAL) tubule segments were dissected and perfused in vitro. The free end of the tubule was held and immobilized at one edge by a holding pipette kept under continuous suction. A micropositioner was used to insert a patch pipette into the lumen, and a gigaohm seal with the luminal membrane was achieved in 455 instances out of considerably more trials. In approximately 20% of all gigaohm seals recordings of single ionic channels were obtained. We have identified only one single type of K⁺ channel in these cell-attached and cell-excised recordings. In the cell-attached configuration with KCl or NaCl in the pipette, the channel had a conductance of 60±6 pS (n=24) and 31±7 pS (n=4) respectively. In cell-free patches with KCl either in the patch pipette or in the bath and with a Ringer-type solution (NaCl) on the opposite side the conductance was 72±4 pS (n=37) at a clamp voltage of 0 mV. The permeability was 0.33±0.02 · 10^±12 cm³/s. The selectivity sequence für this channel was: K⁺=Rb⁺=NH ₄ ⁺ =Cs⁺>Li⁺Na⁺=0; the conductance sequence was K⁺Li⁺Rb⁺=Cs⁺= NH ₄ ⁺ =Na⁺=0. In excised patches Rb⁺, Cs⁺ and NH ₄ ⁺ when present in the bath at 145 mmol/l all inhibited K⁺ currents out of the pipette. The channel kinetics were described by one open (9.5±1.5 ms, n=18) and by two closed (1.4±0.1 and 14±2 ms) time constants. The open probability of this channel was increased by depolarization. The channel open probability was reduced voltage dependently by Ba²⁺ (half maximal inhibition at 0 mV: 0.07 mmol/l) from the cytosolic side. Verapamil, diltiazem, quinine and quinidine inhibited at approximately 1 mol/l ±0.1 mmol/l from either side. Similarly, the amino cations lidocaine, tetraethylammonium and choline inhibited at 10–100 mmol/l. The channel was downregulated in its open probability by cytosolic Ca²⁺ activities > 10^±7 mol/l and by adenosine triphosphate 10^±4 mol/l. The open probability was downregulated by decreasing cytosolic pH (2-fold by a decrease in pH by 0.2 units). The described channel differs in several properties from the K⁺ channels of other epithelia and of renal cells and TAL cells in culture. It appears to be responsible for K⁺ recycling in the TAL segment.Preliminary reports of the present study have been given at the following conferences: Tagung der Deutschen Physiologischen Gesellschaft, Würzburg, October 1988; Membranforum, Frankfurt, April 1989; 3rd Int. Conf. Diur., Mexico City, April 1989; 3rd Nephrology Forefront Symposium, Arrola, July, 1989; IUPS meeting, Helsinki, July 1989. This study has been supported by Deutsche Forschungsgemeinschaft Grant No. Gr 480/9     

Effects of (−)baclofen on inhibitory neurons in the guinea pig hippocampal slice

Intracellular recordings were made from electrophysiologically identified inhibitory neurons in the dentate hilus. (–)Baclofen (0.1–0.5 mol/l), applied by the bath, strongly hyperpolarized inhibitory neurons, reduced their input resistance and induced outward currents under voltage clamp at holding potential of –60 mV in cells recorded with KCl-filled electrodes. Increasing the (–)baclofen concentration (up to 1 mol/l) did not increase the amplitude of the outward current, but increased its duration. (–)Baclofen depressed Cl-dependent IPSPs evoked by perforant path stimulation in inhibitory neurones, granule cells and CA3 neurons. In the case of inhibitory neurons and CA3 neurons, depression of IPSPs, membrane hyperpolarization and increase in membrane conductance concurred. All effects were blocked by BaCl₂ (1 mmol/l) in the superfusate. In the case of granule cells, depression of IPSPs by (–)balcofen out-lasted an only small membrane hyperpolarization, conductance increase or outward current. High concentrations (up to 10 mol/l) of (–)baclofen depressed evoked IPSPs of granule cells for an extended period of time, but the other effects remained small and transient. IPSPs elicited in granule cells by microdrop application of glutamate to the dentate hilus were also blocked by (–)baclofen, but spontaneous IPSPs were only reduced in amplitude. We suggest that the blockade of GABA_A receptor-mediated IPSPs of hippocampal neurons by the GABA_B receptor agonist (–)baclofen can be explained by a K-dependent hyperpolarization of inhibitory neurons.     

Two types of transient outward currents in cardiac ventricular cells of mice

Ventricular cells of adult mice were prepared by an enzyme digestion procedure. Single channel currents were recorded by a conventional patch clamp technique from cell attached patches. Voltage steps from the holding potential of –80 mV to test potentials between –35 and +50 mV caused openings of two types of outward currents through single channels with the conductances of 27 and 12 pS, respectively. The averaged currents reveal transient time courses for both channel types. The current-voltage relations of both single channel currents were linear over the tested voltage range and intersected the voltage axis at –70 mV. This indicates that both single channel currents are mainly carried by potassium ions. All open and closed times were found to be voltage independent. The 27 pS channel had a mean open time of 3.9±1.0 ms (n=8). The closed time consisted of two components with ₁ = 2.1 ± 0.2 ms and ₂ = 50 ± 19 ms (n=8). The 12 pS channel had a mean open time of 34.0±5.2 ms (n=3) and the two components of the mean closed time have been calculated as ₁ = 8.3 ± 2.1 ms and ₂ = 120 ± 50 ms (n=3; all mean ±SD).     

Evidence for Na+/Ca2+ exchange in isolated smooth muscle cells: a fura-2 study

Isolated smooth muscle cells (SMC) from guinea pig taenia coli were employed. Suspension of cells were externally loaded in saline with the fluorescent calcium indicators quin-2/AM or fura-2/AM at 20–40 M or 4 M respectively, resulting in an estimated intracellular concentration of 100–200 M for quin-2 or 10–20 M fura-2 (free acid). On addition of 100 M carbachol or high K _o ⁺ (80 mM) depolarization, fura-2 loaded cells contracted (104±47 m,n=121 rest: 39±13 m,n=59 contracted) identically to control (103±35 m,n=232 rest: 39±16 m,n=89 contracted) cells, whereas quin-2 loaded cells were unresponsive to these protocols and there was no significant length change. The Ca _i ²⁺ of fura-2 loaded cells was 100±18 nM (mean±SD,n=15) and was not significantly different from quin-2 loaded cells 107±26 nM (n=13). Treatment of fura-2 loaded cells with 100 M ouabain saline for 10–60 min progressively elevated the Ca _i ²⁺ to a mean of 266±83 nM (n=15). Reduction of Na _p ⁺ (96% Li⁺ replaced) significantly increased Ca _i ²⁺ to 317±77 nM (n=8). After pretreatment with ouabain (100 M), Na _o ⁺ replacement (Li⁺) increased Ca _i ²⁺ at a significantly faster rate [3.6 nM min^–1 (control) cf. 19.8 nM min^–1 (ouabain)].