Phorbol 12,13-dibutyrate,an activator of protein kinase C,stimulates both contraction and Ca2+ fluxes in dog saphenous vein

Summary The vascular effects of phorbol 12,13-dibutyrate (PDBu) were studied in the dog saphenous vein. PDBu (1 M) caused contraction (0.58 ± 0.22 g/mg wet wt.) and Ca uptake (74.2 ± 41.2 mol/kg wet wt.) which were unaffected by 10 M phentolamine (N = 6). The PDBu-induced contraction was greatly (60–80%) inhibited in Ca²⁺-free solution. 15 Ca efflux measurements performed in Ca²⁺-free solution showed that PDBu did not cause Ca release from intracellular storage sites. The contractile response to PDBu (1 nM-1 M) was significantly correlated with the magnitude of Ca uptake; contraction and the rise in tissue Ca²⁺ also had a similar time course. Correlation between the two measures persisted when the responses to PDBu were augmented by co-administration with 20 mM KCl. However, no synergism occurred between the two agonists. Both the contraction and Ca uptake responses to PDBu were reduced by nifedipine and verapamil, each at 1 M. In the Triton X-100 skinned saphenous vein, where the voltage-dependent Ca channel is not functional, 10 M PDBu did not cause contractions in the presence of 0.1 M Ca²⁺. Thus, contraction of the intact saphenous vein by PDBu characteristically exhibits great Ca dependence and PDBu seems to activate the voltage-dependent Ca channel, presumably through stimulation of protein kinase C; the ensuing Ca entry is primarily responsible for contraction. However, the mechanism responsible for the PDBu-induced contractions that are resistant to Ca²⁺-free PSS or Ca entry blockers remains to be defined. It appears that the dog saphenous vein differs from dog femoral artery, rabbit aorta and pig carotid artery where PDBu contractions do not display dependence on external Ca²⁺. Send offprint requests to P. J. S. Chin at the above address     

Phenytoin partially antagonized L-type Ca2+ current in glucagon-secreting tumor cells (ITC-1)

Summary Transmembrane Ca²⁺ currents were investigated by means of a whole-cell clamp technique in a hamster glucagon-secreting tumor cell line (ITC-1). Two types of Ca²⁺ current were identified in ITC-1 cells. The low-threshold and transient (T-type) current became detectable above the potential level around –60 mV and decayed rapidly with an inactivation time constant of 95 ms (at –40 mV and 23°C), while the high-threshold and long-lasting (L-type) one was activated by depolarization more positive to –30 mV with non-inactivating kinetics. The voltage dependence and kinetics of these currents were identical to those reported in guinea-pig pancreatic ₂ cells. Both currents were augmented by equimolar substitution of Ca²⁺ with Ba²⁺ and completely abolished by adding 1 M La³⁺. Phenytoin, a well known anti-epileptic drug and a postulated T-type specific Ca²⁺ current antagonist, surprisingly blocked the L-type current without affecting the T-type current in ITC-1 cells. While phenytoin antagonized the L-type Ba²⁺ current selectively, 60% of the current remained even in supramaximal concentration range over 500 M. The residual component of the L-type current was completely abolished by adding nifedipine.     

Steady-state concentrations of choline and acetylcholine in rat brain parts during a constant rate infusion of deuterated choline

Summary An intravenous infusion of deuterated choline at constant rate for 6 min (5 or 25 moles kg^–1 min^–1) significantly increases the concentration of choline in plasma, occipital cortex and striatum. Both 5 and 25 moles kg^–1 min^–1 increase the concentration of acetylcholine in cortex but only 25 moles kg^–1 min^–1 increases the acetylcholine content in striatum. In contrast, 1 mole kg^–1 min^–1 does not change the choline or acetylcholine content in cortex or striatum. A single pulse injection of choline (200 moles kg^–1) causes a significant increase in the concentration of choline in striatum 30 sec following injection. The choline content returns to normal values within 2 min. These studies show that when a pulse injection of a non-tracer dose of radioactive choline is used to measure brain acetylcholine turnover rate the maintenance of steady state must be verified within seconds after the pulse injection of radioactive choline. When constant infusion of deuterated choline is used to measure turnover rate of acetylcholine in the brain of rats, a dose of 1 mole kg^–1 min^–1 appears to be a maximal infusion rate.     

Effects of lead on cloned voltage-operated neuronal potassium channels

The action of lead (Pb ²⁺) on cloned voltage-operated potassium channels of the rat brain was investigated in oocytes of Xenopus laevis. Pb²⁺ was found to decrease the potassium currents. This effect was due to a shift of the current-voltage relation in a positive direction (up to 30 mV). The Pb²⁺ effect appeared at a threshold concentration of about 0.1 mol/l and was maximal at a concentration of about 30 mol/l. At a potential of – 30 mV, the concentration needed for a 50% reduction of the potassium current was 1.0 mol/l. The depressant effect of Pb²⁺ was obtained with all potassium channels tested (Kv1.1, Kv1.2, Kv1.4, Kv2.1, Kv3.4). It was minimal for the Kv2.1 channel and maximal for the Kv1.1 channel at potentials negative to 0 mV. An effect comparable with that of Pb²⁺ could not be induced by the application of magnesium or calcium. The external application of Pb²⁺ led to a decrease of potassium currents in outside-out but not in inside-out membrane patches. Overall, Pb²⁺ had a significant effect on the potassium channels which may contribute to the mechanisms of Pb²⁺ neurotoxicity.     

The Potential Use of the South African River Crab,Potamonautes warreni,as a Bioindicator Species for Heavy Metal Contamination

In 1995, preliminary water and sediment analyses of the river bed and burrow sediments from 9 locations along the Mooi River, NW Province, South Africa had shown cadmium concentrations up to 0.009 mg l^–1±0.003 and up to 0.33 and 0.89 weight % with scanning electron microscopy (SEM) and x-ray microanalysis. Samples of the adult river crab (Potamonautes warreni) were collected from the Mooi River at Noordbrug (26°40S/27°05E), 1 km north of Potchefstroom Town, and exposed to 0.2 or 2.0 mg Cd²⁺ l^–1 in situ to determine tolerance, uptake and bioaccumulation of cadmium. Using flame atomic absorption spectroscopy (FAAS) the gills, haemolymph and digestive gland of naturally exposed P. warreni showed wet mass values of 0.74±0.27 g Cd²⁺ g^–1, 0.007±0.007 g ml^–1 and 0.12±0.09 g g^–1 respectively. The tolerance of crabs to aqueous Cd reached its limit (ET₅₀=42 hours) at 2.0 mg l^–1 aqueous Cd exposure. At an exposure to 0.2 mg Cd²⁺ l^–1 for 21 days, the greatest Cd (n=11; 9.99±5.09 g g^–1 wet mass) and Cu concentrations (n=11; 17.90±4.66 g g^–1 wet mass) were associated with the gills, and to a lesser extent the digestive gland (n=11; 0.38±0.20 g g^–1 wet mass), whereas concentrations of Zn were variable in both organs. In the haemolymph Cd levels were relatively small (n=11; 0.012–0.006 g ml^–1) with exposure and time and Cu, Zn concentrations varied. Changes in the uptake of Cd in P. warreni indicated that transport, storage and possibly regulatory mechanisms are likely to operate in adult crabs. The potential of P. warreni as a bioindicator species of pollution is also discussed.     

Characteristic behavioral alterations in rats induced by rolipram and other selective adenosine cyclic 3′,5′-monophosphate phosphodiesterase inhibitors

The significance of a characteristic symptomatology (hypothermia, hypoactivity, forepaw shaking, grooming, head twitches) as a potential in vivo correlate of enhanced availability of brain adenosine cyclic 3,5-monophosphate (cAMP) was examined in rats following systemic administration of various doses of dibutyryladenosine cAMP (dBcAMP) or of the phosphodiesterase (PDE) inhibitors rolipram, Ro 20-1724, ICI 63-197, isobutylmethylxanthine (IBMX), theophylline, cartazolate, and papaverine. The various PDE inhibitors could be assigned to three groups according to the pattern of behavioral alterations they induced. Rolipram, Ro 20-1724, and ICI 63-197 (group 1) caused hypothermia, hypoactivity, forepaw shaking, grooming, and head twitches. All behavioral effects were mimicked by dBcAMP but not dBcGMP. The order of potency and effective dosage range to induce the behavioral alterations were, in descending order, rolipram (0.09–1453 mol/kg IP), ICI 63-197 (0.48–119 mol/kg IP), Ro 20-1724 (5.6-1438 mol/kg IP), corresponding with the recently reported efficacy of the drugs to elevate rat brain cAMP in vivo. Comparatively high doses of the alkylxanthine PDE inhibitors IBMX and theophylline (group 2) caused hypothermia, forepaw shaking, grooming, and head twitches concomitantly with a decline of the motor stimulatory effect, suggesting enhanced availability of brain cAMP. The order of potency and the effective dosage range to induce the behavioral alterations were, in descending order, IBMX (28.1–113 mol/kg IP) and theophylline (139–555 mol/kg IP). The third group, papaverine (295–1179 mol/kg IP) and cartazolate (21.5–345 mol/kg IP), caused only hypothermia and hypoactivity. The differences in the behavioral pattern of the two latter groups of compounds in comparison with dBcAMP and the selective cAMP PDE inhibitors are discussed with regard to their additional interference with adenosine actions besides their nonselective PDE inhibitory action.With the technical assistance of Petra Zehleke     

Effects of saterinone and its enantiomers R(+)-saterinone and S(−)-saterinone on the phosphodiesterase isoenzymes from ventricular tissue of failing human hearts and porcine hearts

Summary Stereoisomers often exhibit differences in their pharmacological activities. Therefore, the phosphodiesterase inhibitory effects of the cardiotonic agent saterinone in form of the racemate were investigated in comparison with the inhibitory properties of its enantiomers R(+)- and S(–)-saterinone. For this purpose the phosphodiesterase isoenzymes from ventricular tissue of failing human hearts and porcine hearts were separated by DEAE-sepharose anion exchange chromatography. Four different phosphodiesterase isoenzymes were isolated from failing human myocardium and designated phosphodiesterase I–IV. Three phosphodiesterase isoenzymes could be separated from ventricular tissue of porcine hearts. A Ca²⁺/calmodulin stimulated phosphodiesterase I was not detectable in porcine myocardium.In failing human hearts racemic saterinone was a potent inhibitor of phosphodiesterase III (IC₅₀ 0.02 mol/l) and IV (IC₅₀ 0.03 mol/l) as compared to the inhibition of phosphodiesterase I (IC₅₀ 37.3 mol/l) and II (IC₅₀ 51.4 mol/l). In comparison with the racemate, R(+)- and S(–)-saterinone showed only slight differences in their phosphodiesterase inhibitory effects. R(+)-saterinone inhibited phosphodiesterase III slightly but significantly more potently and selectively than did S(–)-saterinone. Compared to the inhibition of phosphodiesterase I and II both enantiomers were similarly potent and selective inhibitors of phosphodiesterase III and IV. Similar results were obtained in porcine hearts.It is concluded that the racemate saterinone and its enantiomers R(+)- and S(–)-saterinone are virtually equipotent concerning the inhibition of phosphodiesterase isoenzymes isolated from failing human hearts or porcine ventricular tissue. The enantiomers of saterinone did not exhibit distinct stereoselectivity in their phosphodiesterase inhibitory effects.Parts of the results have been presented at the Autumn Meeting in Berlin (Zimmermann et al. 1991) and at the 33th Spring Meeting in Mainz (Wenzlaff et al. 1992) of the Deutsche Gesellschaft für Pharmakologie und Toxikologie Correspondence to: W. Zimmermann at the above address     

Characterisation of the mechanisms for hyperactivity induction from the nucleus accumbens by phenylethylamine derivatives

A wide variety of phenylethylamine derivatives were injected bilaterally into the nucleus accumbens of rat following a nialamide pretreatment and hyperactivity was recorded. 2-Phenylethylamine was shown to induced a low intensity hyperactivity but the inrroduction of hydroxyl functions on to the phenyl ring at the 3- and/or 4-positions enhanced activity and m- and p-tyramine and dopamine each caused marked hyperactivity in the 0.4–25 g dosage range. Methylation of one hydroxyl function reduced activity (3-methoxy-4-hydroxy- and 3-hydroxy-4-methoxy-phenylethylamine); 2(3,4-methylenedioxyphenyl) ethylamine was inactive. Agents with substitution of the side chain, such as noradrenaline, d-amphetamine and -methyldopamine, were all shown to induced marked hyperactivity at doses of 1.6–25 g. Alterations in the chain length markedly reduced activity (4-(3,4-dihydroxyphenyl) butylamine, 3,4-dihydroxybenzylamine). A variety of N-substituted compounds were shown to be potent inducers of hyperactivity from the nucleus accumbens (adrenaline, epinine, N-ethyldopamine, N-isopropyldopamine, isoprenaline) (0.2–25 g). However, N-methyl-N-isopropyldopamine showed only weak activity and N,N-dimethyldopamine was inactive. All hyperactivity effects were shown to be dose-dependent. The hyperactivities induced by dopamine, noradrenaline and isoprenaline were each inhibited in a dose-dependent manner by subsequent injections of fluphenazine (1.25–25 g) into the nucleus accumbens, although no reductions were recorded following similar injections of saline, solvent, 2% procaine, 50 g propranolol or 50 g piperoxan.     

Comparative toxicity of fostriecin,hepsulfam and pyrazine diazohydroxide to human and murine hematopoietic progenitor cells in vitro

Summary The in vitro myelotoxic potentials of three investigational antitumor agents, Fostriecin, Hepsulfam and pyrazine diazohydroxide (PZDH), were evaluated utilizing clonogenic assays. Human and murine marrow cells were exposed to each drug for 1 hr prior to culture in microcapillary (human) or Petri dish (murine) assays. Fostriecin (0.22–220 M), Hepsulfam (0.34–340 M) and PZDH (0.68–680 M) inhibited myeloid (CFU-gm), erythroid (BFU-e, CFU-e) and megakaryocytic (CFU-meg) colony formation in a concentration-dependent manner. CFU-e from both species were more sensitive to Fostriecin than the other progenitors and murine cells more sensitive overall to Fostriecin than their human counterparts. Murine CFU-e were also more sensitive to Hepsulfam than human CFU-e, with CFU-gm and BFU-e being similarly affected in both species. Human BFU-e were greatly inhibited by PZDH, whereas murine BFU-e were relatively resistant to its toxic effects. Fostriecin was the most toxic of the three antitumor agents, with PZDH the least toxic.     

Developing gossypol derivatives with enhanced antitumor activity

Summary Preclinical and clinical studies have pointed to the antitumor potential of the naturally occurring polyphenolic binaphthyl dialdehyde, gossypol, as well as its purified (–,+) enantiomers. To explore further the antitumor properties of this multifunctional agent, we synthesized several reactive derivatives including the (–,+) enantiomers of gossypolone and four different gossypol Schiffs bases (AR1, AR2, AR3, AR4). The biological activities of these new agents were screened by measuring theirin vitro antiproliferative activity against malignant (MCF-7, MCF-7/adr) or immortalized (HBL-100) human breast epithelial cell lines. Racemic gossypolone showed relatively uniform antiproliferative activity against all of the breast epithelial cell lines with 3- to 5-fold less activity than (-)-gossypol against MCF-7 and MCF-7/adr cells. Of interest, the relative antitumor potency of purified gossypolone enantiomers was reverse that of gossypol enantiomers, since (+)-gossypolone showed up to 3-fold greater inhibition of MCF-7 culture growth than (-)-gossypolone. Of the Schiff's base derivatives only AR3 with its isopropyl amine substituent demonstrated cytotoxic activity comparable to that of (-)-gossypol; derivatives with ethyl, propyl, or butyl amine substituents (AR1, AR2, AR4) had little growth inhibitory activity at culture concentrations up to 25 M. AR3 activity was greatest against HBL-100 and MCF-7 cells [MCF-7 IC₅₀ values: AR3=0.9 M, (-)-gossypol=2.3 M]; unlike (-)-gossypol, however, AR3 showed substantially reduced activity against the multidrug-resistant subline, MCF-7/adr. These structure-activity comparisons suggest that isolation of (–,+)-enantiomers of AR3 and additional chemical modifications including the synthesis of an isopropyl amine Schiff's base of gossypolone will likely yield a newer generation of gossypol analogues with enhanced anticancer potential.     

An electrophysiological investigation of the effects of cisplatin and the protective actions of dexamethasone on cultured dorsal root ganglion neurones from neonatal rats

It this study we have investigated the acute and chronic effects of cisplatin on whole cell currents in cultured dorsal root ganglion neurones. Consistent with effects on action potentials measured under current clamp, acute (5 min) application of cisplatin (5 M) attenuated voltage-activated potassium, and mixed cation currents by approximately 50% in both cases. Chronic treatment (5–7 days) of cultured neurones with 5 M cisplatin also resulted in greatly reduced voltage-activated potassium currents (by 50%) and calcium currents (by 60%) compared to events recorded from neurones not treated with cisplatin. In contrast, the amplitude of inward cation current activated by hyperpolarization was doubled by 5–12 days treatment with cisplatin. Studies on action potential after-depolarizations and calcium-activated chloride currents suggest that cisplatin disturbs calcium homeostatic mechanisms. These observations may account for anode break spike excitation and the low efficiency with which cells buffer intracellular calcium following cisplatin treatment.Dexamethasone has been found to enhance the anti-emetic effects of 5-HT₃ receptor antagonists in patients treated with cisplatin. For this reason the actions of dexamethasone were studied in combination with cisplatin treatment. Although acute application of dexamethasone (1–10 M) produced transient depolarizations and bursts of action potentials, after 5 minutes application it had no effect on membrane potential, input resistance, or the properties of action potentials evoked by depolarizing current commands. Compared to cells exposed to cisplatin alone, combined treatment of cisplatin and dexamethasone significantly improved survival of dorsal root ganglion neurones in culture by 20%. Dexamethasone also showed signs of protecting neurones from cisplatin by improving membrane potentials and action potential thresholds. In conclusion, cisplatin reduces the viability of dorsal root ganglion neurones in culture and alters their electrophysiological properties. Evidence suggests that dexamethasone has some protective properties against the neurotoxic actions of cisplatin.     

Inhibitory effects of propiverine on rat and guinea-pig urinary bladder muscle

Summary In muscle strips isolated from guinea-pig and rat urinary bladder, propiverine (3–10 M) inhibited carbachol-induced contractions in the presence of verapamil and Ca²⁺-induced contractions in excess K⁺ medium containing atropine, suggesting it has both anticholinergic and Ca²⁺ channel blocking actions.The Ca²⁺ channel blocking action was also demonstrated by recording inward Ca ²⁺ currents in single cells dispersed from both species. The inhibition of inward currents by propiverine was three times stronger in the rat than the guinea-pig, ID₅₀ being 7 M for rat and 21 M for guinea-pig. The recovery of the current after washout was faster than that of mechanical inhibition. It is concluded that propiverine blocks not only muscarinic receptors, but also Ca²⁺ channels at similar concentrations.Correspondence to: T. Tomita at the above address     

Modulation of calcium-currents by capsaicin in a subpopulation of sensory neurones of guinea pig

Summary The action of capsaicin (CAP) on the total Ca²⁺ current was examined in internally perfused voltage-clamped dorsal root ganglion (DRG) neurones of guinea pigs. CAP changed the total Ca²⁺ current in about 50% of the investigated DRG neurones (CAP-sensitive neurones) in the following way: (I) a transient increase of the current amplitude at potentials between – 35 mV and about – 10 mV was accompanied by a shift of the current-voltage relation towards negative potentials by 5–8 mV; (II) the current inactivation was accelerated at potentials positive to about – 35 mV; and (III) the current activation of Ca²⁺ currents (time to peak values) was also accelerated. Separated low voltage-activated (T-type) currents at potentials negative to about – 35 mV were either not affected or reduced. It remains undecided whether CAP increases T-type currents in a particular potential range or activates an N-type current. External application of 50 M Ni²⁺ blocks the effect of CAP, but does not affect the acceleration of the high voltage-activated (L-type) current inactivation induced by menthol. This appears to exclude a CAP effect on L-type current inactivation. CAP sensitive and CAP insensitive neurones could be discriminated by their different Ca²⁺ currents: the former demonstrate both fast and slow inactivating currents while the latter have only L-type currents. The observed changes of fast-inactivating Cat²⁺ currents may be related to the specific action of CAP on peptidergic sensory neurones. Send offprint requests to M. Petersen     

Triethyllead in the biological material in cases of acute tetraethyllead poisoning

Summary Triethyllead and the total lead were determind in the tissues in three acute fatal cases of tetraethyllead poisoning. In each case high concentrations of triethyllead were found (2.0–22.0 g/g) being of the same range as the total lead content. The triethyllead content of the tissues showed a slightly decreasing trend with the length of the surviving period. High levels of the total lead were also found in blood (3.3–4.0 g/ml), and urine (0.4–8.0 g/ml), the highest values being observed after EDTA treatment. The triethyllead content in blood and urine was much lower.     

Actions of the phosphodiesterase inhibitor zardaverine on guinea-pig ventricular muscle

Summary The positive inotropic action of the phosphodiesterase inhibitor zardaverine was investigated in guinea-pig heart muscle. In right papillary muscles, 1–30 M zardaverine reversibly increased the force of contraction in a concentration-dependent manner. This effect was accompanied by a shortening of contraction and relaxation times. Resting membrane potential was unchanged, whereas action potential amplitude was significantly increased and duration was reduced. In papillary muscles partially depolarised with 22 mM K⁺, zardaverine (10 and 30 LM) restored slow action potentials, which were not influenced by cimetidine, propranolol or prazosin but were blocked by the calcium channel blocker (+)-nitrendipine or the muscarinic agonist carbachol. cAMP-specific phosphodiesterase III, isolated from guinea-pig ventricular muscle was inhibited by zardaverine as was cAMP-specific phosphodiesterase IV, isolated from dog trachea (IC₅₀s: 0.5 and 0.8 M, respectively). The results suggest that the observed positive inotropic and electrophysiological effects result from an inhibition of cellular phosphodiesterase.Send offprint requests to C. Schudt at the above address     

Inhibition of potassium outward currents and pacemaker current in sheep cardiac Purkinje fibres by the verapamil derivative YS 035

Summary The electrophysiologic mode of action and potency of the verapamil derivative YS 035 (N,N-bis-(3,4-dimethoxyphenethyl)-N-methyl amine) were investigated in sheep cardiac Purkinje fibres. Action potential duration measured at a repolarization level of –60 mV (APD-60) and membrane currents recorded with the two-microelectrode voltage-clamp technique were evaluated. At 10 mol/l YS 035 APD-60 was increased to about 115% of reference. Prolongation measured as percentage of the respective control exhibited on the average no dependence on stimulation frequency (0.17–2 Hz). At 100 mol/l membrane became depolarized to about –50 mV and action potentials could no longer be elicited. Further study was focussed on effects on outward currents, mostly activated at a frequency of 0.05 Hz. Transient outward current (i_to) was completely blocked at 100 mol/l and half-maximal inhibition occurred at about 14 mol/l. Inwardly rectifying potassium current (i_K1) was reduced to 47% of reference at 100 mol/l. An initially activating outward current at positive membrane potentials (i_inst) was reduced to 73% at 100 mol/l. Time-dependent (delayed) outward current (i_K) was on the average not affected up to 100 ol/l. Besides inhibition of repolarizing outward currents YS 035 completely blocked pacemaker current (i_f) at 100 mol/l and half-maximal reduction was achieved at 5 mol/l. YS 035 (1–100 mol/l) did not clearly affect time constants of activation at selected test potentials (I_K: +35 mV; i_f: –90 mV) or inactivation (i_to: 0 mV). Voltage-dependent control mechanisms of currents (it_to, i_f) were not influenced by YS 035 but the amount of available current was reduced.In conclusion, the verapamil derivative YS 035 inhibited pacemaker current and potassium outward currents which correlated to a prolongation of cardiac action po tentials. Electrophysiological actions of the compound favour it to be tested in vivo as an antiarrhythmic drug candidate. Send offprint requests to U. Borchard at the above address     

The low KM-phosphodiesterase inhibitor denbufylline enhances neuronal excitability in guinea pig hippocampus in vitro

Summary The actions of the phosphodiesterase inhibitor denbufylline on the excitability of hippocampal neurons were investigated by means of extracellular and intracellular recordings. Denbufylline, which has been shown to selectively inhibit a low K_M, Ca²⁺/calmodulin-independent phosphodiesterase isozyme, concentration-dependently increased the amplitude of the extracellularly recorded CAI population spike evoked by electrical stimulation of the Schaffer collateral/commissural pathway. Concentration-response-curves yielded an EC₅₀ for denbufylline of 0.76 M. In comparison, the nonselective phosphodiesterase inhibitor 3-isobutyl-lmethylxanthine (IBMX) also produced an increase in the amplitude of the population spike. From the concentration-response-curve, which was steeper than that of denbufylline, an EC₅₀ for IBMX of 1.04 M was obtained. However, despite their similar EC₅₀ values, denbufylline was found to be significantly more potent at lower concentrations (<- 300 nM) than IBMX.Intracellular recordings from CAI pyramidal cells revealed postsynaptic actions of denbufylline (300 nM) as indicated by a small drug-induced depolarization (2 – 5 mV) associated with an increase in membrane input resistance by 10–20%. In addition, denbufylline blocked the accommodation of trains of action potentials evoked by the injection of depolarizing current pulses.The results suggest i) that accumulation of adenosine-3,5-monophosphate (CAMP) in the postsynaptic cell and/or in the presynaptic terminal produced by blockade of phosphodiesterases leads to enhanced synaptic transmission in the CAI area of the hippocampus and ii) that a low K_M, Ca ²⁺/calmodulin-independent cAMP-phosphodiesterase is an important component involved in the regulation of the intracellular cAMP level at synapses of central nervous system neurons. Send offprint requests to B. Sutor at the above address     

Abolition by calcium antagonists of the autoregulation of renal blood flow

Summary Intra-arterial infusions of Ca antagonistic vasodilator substances such as nifedipine (1–10 g/min) and verapamil (30–100 g/min) inhibited renal autoregulation in the perfused dog kidney. Simultaneous infusion of 30 mg/min of CaCl₂ with nifedipine (3 g/min) or with verapamil (30 g/min) abolished the effects of these Ca antagonists. It is suggested that Ca ions play an important role in establishing renal autoregulation.     

Effects of the calmodulin antagonists fendiline and calmidazolium on aggregation,secretion of ATP,and internal calcium in washed human platelets

Summary Ca²⁺-calmodulin dependent phosphorylation of myosin is essential for the induction of platelet shape change and subsequent reactions. Therefore, we studied the effects of the calmodulin antagonists fendiline and calmidazolium on the thrombin-induced aggregation, secretion of ATP, and increases in the intracellular free calcium concentration ([Ca²⁺]_i) in washed human platelets in the absence and presence of extracellular Ca²⁺. In Ca²⁺ free medium, fendiline (10–100 M) and calmidazolium (3–30 M) concentration-dependently inhibited aggregation. The effect of fendiline could be partly reversed by extracellular Ca²⁺ and higher thrombin concentrations. Furthermore, aggregations induced by the calcium ionophore ionomycin and by the protein kinase C-activator 4--phorbol 12-myristate 13-acetate were inhibited by fendiline, although to a smaller degree than the thrombin-induced aggregation. Thrombin-induced secretion of ATP was attenuated by low concentrations of fendiline (1–30 M) and calmidazolium (1 M) but enhanced by higher concentrations (10–30 and 3–10 M, respectively), independently of extracellular Ca²⁺. Fendiline (1–10 M) did not affect [Ca²⁺]_i in resting and thrombin-stimulated platelets. At higher concentrations (30–100 M), it induced increases in [Ca²⁺]_i in unstimulated platelets and attenuated the response to thrombin in Ca²⁺ free medium, whereas thrombin-induced Ca²⁺ influx was markedly enhanced. Similar results were obtained with calmidazolium (1–3 M). These stimulating effects on ATP secretion and on [Ca²⁺]_i of fendiline and calmidazolium may be attributed to interactions with platelet membranes by which the permeability of small cations is increased. In contrast to these actions that would be normally considered platelet-activating, our study demonstrates that the two calmodulin antagonists effectively antagonize [Ca²⁺]_i ^– mediated induction of platelet aggregation. Send offprint requests to A. Lückhoff at the above address     

Effects of (+)- and (±)-sotalol on repolarizing outward currents and pacemaker current in sheep cardiac Purkinje fibres

Summary This study was aimed to differentiate the action of (+)- and (±)-sotalol (10–1000 mol/l) on membrane currents which are active during the repolarization of cardiac action potentials Effects where studied in shortened sheep cardiac Purkinje fibres with the two-microelectrode voltage-clamp technique Action potentials were activated at a frequency of 0.25 Hz and membrane currents at 0.03 Hz or 0.05 Hz in most experiments.Out of the currents investigated the transient outward current (i_to) reacted most sensitively to (+)- and (±)-sotalol. I_to-amplitude was decreased on the average to 77% of reference at 10 mol/l and to 53% at 1000 mol/l (+)- or (±)-sotalol. The maximally available ito-current was decreased but the voltage-dependent control of inactivation was left nearly unchanged. The initial inwardly rectifying current (i_Ki), which propels the last repolarization phase of the action potential and controls resting potential to a large extent was reduced on the average to 93% of reference at 10 mol/l and to 62% at 1000 mol/l (+)- or (±)-sotalol. Time-dependent (delayed) outward current (i_K) was on the average not affected by (+)- or (±)-sotalol up to 100 mol/l and was decreased to 84% of reference current under the influence of 1000 mol/l. An initial outward current, which is activated at positive membrane potentials (i_inst) was not clearly affected by (+)- or (±)-sotalol at concentrations up to 1000 mol/l Pacemaker current (i_f) was not influenced by the drugs up to 100 mol/l. Only at 1000 mol/l was the amount of available i_f-current decreased to 79% of reference. (The potential-dependent control of activation was not affected) Time constants of time-dependent currents i_to, i_K and i_f did not change in concentrations up to 1000 mol/l of the drug.Action potential duration increased at (+)- or (±)-sotalol concentrations 10 mol/l and maximal prolongation was achieved at concentrations of 100–300 mol/l Resting potential remained nearly unchanged at these concentrations, but the membranes depolarized at 1000 mol/l. According to our data action potential prolongation in sheep Purkinje fibres under the influence of (+)- and (±)-sotalol correlates to the drug-induced block to i_to-current and inwardly rectifying i_K1-current.Supported by the Deutsche Forschungsgemeinschaft SFB 242, C 1 Send offprint requests to U. Borchard at the above address