Graft copolymerization of methyl methacrylate on chlorinated natural rubber, 3. Study of the interaction between chlorinated and organometallic compounds

The interaction of Ni[P(OR)₃]₄ (R = C₂H₅ or C₆H₅) with chlorinated rubber (CR) of CCl₄ is studied. The results of the polymerization of methyl methacrylate (MMA) initiated by the system CR or CCl₄? Ni[P(OR)₃]₄ depend on the order in which the reagents are introduced. Trialkylphosphite, tetrakis(trialkylphosphite) nickel and the products resulting from the reaction of CCl₄ or CR with Ni[P(OR)₃]₄ are characterized by ³¹P NMR. From this study, it appears that the initiation of polymerization by the system chlorinated compound-nickel derivative does not involve the formation of free phosphite, which is at variance with previously reported assumption. ¹H NMR study supports these results.     

Agonist-induced calcium flux,phosphoinositide metabolism,aggregation and enzyme secretion in human neutrophils

Formyl-methionyl-leucyl-phenylalanine (FMLP), platelet activating factor (PAF) and leukotriene B₄ (LTB₄) are potent activators of human neutrophils. Using human neutrophils prelabelled with the fluorescent indicator dye, Quin 2, or with [³²P]-orthophosphate, we examined the effects of these stimuli on intracellular free calcium concentration, [Ca²⁺]i, and, on various indices of phosphoinositide metabolism, including [³²P]-phosphatidic acid (PtdA) formation. The concentration-dependence of the observed changes in [Ca²⁺]i or [³²P]-PtdA were then compared to stimulus-induced aggregation and enzyme release (-N-acetylglucosaminidase (NAG) and lysozyme).FMLP, PAF and LTB₄ caused a concentration-dependent elevation of [Ca²⁺]i, aggregation and enzyme release. However, unlike FMLP and PAF, LTB₄ (2.5 M) did not cause significant formation of [³²P]-PtdA. The concentration response curves for agonist-induced elevation of [Ca²⁺]i lie to the left of those for aggregation and enzyme release. FMLP and PAF also caused an elevation of [Ca²⁺]i at concentrations lower than those required to elicit [³²P]-PtdA formation.These observations suggest that [Ca²⁺]i elevationper se cannot mediate human neutrophil functional responses to FMLP, PAF and LTB₄. Consequently there may exist other mediator(s) that act in concert with [Ca²⁺]i or are triggered by [Ca²⁺]i elevation to promote human neutrophil activation. Both the elevation of [Ca²⁺]i and the formation of these putative mediator(s) in response to LTB₄ apparently occur independently of inositol phospholipid hydrolysis.     

Contribution of pH,diprotonated phosphate and potassium for the reflex increase in blood pressure during handgrip

The relative importance of pH, diprotonated phosphate (H₂PO₄^?) and potassium (K⁺) for the reflex increase in mean arterial pressure (MAP) during exercise was evaluated in seven subjects during rhythmic handgrip at 15 and 30% maximal voluntary contraction (MVC), followed by post-exercise muscle ischaemia (PEMI). During 15% MVC, MAP rose from 92 ± 1 to 103 ± 2 mmHg, [K⁺] from 4.1 ± 0.1 to 5.1 ± 0.1 mmol L^?1, while the intracellular (7.00 ± 0.01 to 6.80 ± 0.06) and venous pH fell (7.39 ± 0.01 to 7.30 ± 0.01) (P < 0.05). The intracellular [H₂PO₄^?] increased 8.4 ± 2 mmol kg^?1 and the venous [H₂PO₄^?] from 0.14 ± 0.01 to 0.16 ± 0.01 mmol L^?1 (P < 0.05). During PEMI, MAP remained elevated along with the intracellular [H₂PO₄^?] as well as a low intracellular and venous pH. However, venous [K⁺] and [H₂PO₄^?] returned to the level at rest. During 30% MVC handgrip, MAP rose to 130 ± 3 mmHg, [K⁺] to 5.8 ± 0.2 mmol L^?1, the intracellular and extracellular [H₂PO₄^?] by 20 ± 5 mmol kg^?1 and to 0.20 ± 0.02 mmol L^?1, respectively, while the intracellular (6.33 ± 0.06) and venous pH fell (7.23 ± 0.02) (P < 0.05). During post-exercise muscle ischaemia all variables remained close to the exercise levels. Analysis of each variable as a predictor of blood pressure indicated that only the intracellular pH and diprotonated phosphate were linked to the reflex elevation of blood pressure during handgrip.     

1H and 13C NMR sturctural investigation of some titanium alkoxide-magnesium halide complexes as Ziegler-Natta catalyst precursors

Several [Ti₂(OR)₈MtX₂]₂ complexes were prepared from MtX₂ (Mt = Mg, Zn; X = Cl, Br, I) and Ti(OR)₄ (R = ethyl, propyl, butyl). ¹H and ¹³C NMR in toluene-d₈ solution and cross-polarization/magic-angle spinning (CP/MAS) ¹³C NMR spectra were taken of some complexes. The obtained spectra allowed to deduce their molecular structure in solution and to compare them with that of [Ti₂(OC₂H₅) ₈Cl]₂Mg₂Cl₂ obtained by single-crystal X-ray diffraction analysis.     

Na+-dependent regulation of the free Mg2+ concentration in neuropile glial cells and P neurones of the leech Hirudo medicinalis

 To investigate the Mg²⁺ regulation in neuropile glial (NG) cells and pressure (P) neurones of the leech Hirudo medicinalis the intracellular free Mg²⁺ ([Mg²⁺]_i) and Na⁺ ([Na⁺]_i) concentrations, as well as the membrane potential (E _m), were measured using Mg²⁺- and Na⁺-selective microelectrodes. The mean steady-state values of [Mg²⁺]_i were found to be 0.91 mM (mean E _m=–63.6 mV) in NG cells and 0.20 mM (mean E _m=–40.6 mV) in P neurones with a [Na⁺]_i of 6.92 mM (mean E _m=–61.6 mV) and 7.76 mM (mean E _m=–38.5 mV), respectively. When the extracellular Mg²⁺ concentration ([Mg²⁺]_o) was elevated, [Mg²⁺]_i in P neurones increased within 5–20 min whereas in NG cells a [Mg²⁺]_i increase occurred only after long-term exposure (6 h). After [Mg²⁺]_o was reduced back to 1 mM, a reduction of the extracellular Na⁺ concentration ([Na⁺]_o) decreased the inwardly directed Na⁺ gradient and reduced the rate of Mg²⁺ extrusion considerably in both NG cells and P neurones. In P neurones Mg²⁺ extrusion was reduced to 15.4% in Na⁺-free solutions and to 6.0% in the presence of 2 mM amiloride. Mg²⁺ extrusion from NG cells was reduced to 6.2% in Na⁺-free solutions. The results suggest that the major [Mg²⁺]_i-regulating mechanism in both cell types is Na⁺/ Mg²⁺ antiport. In P neurones a second, Na⁺-independent Mg²⁺ extrusion system may exist. Received: 11 August 1998 / Received after revision: 14 October 1998 / Accepted: 15 October 1998     

Interaction of polystyryl radical with copper(II) complexes

The polymerization of styrene initiated by 2,2′-azoisobutyronitrile (AIBN) was studied in N,N-dimethylformamide (DMF) solution at 60°C in the presence of tetrakis(N,N-dimethylformamide)copper(II) perchlorate, and also in the presence of its monoazido copper(II) complex [Cu(DMF)₃N₃]⁺. The monoazido complex in DMF was prepared in situ by mixing solid sodium azide with tetrakis(N,N-dimethylformamide)copper(II) perchlorate in a mole ratio of 1:1. The nature of the complex was established by Job's method. The equilibrium constant K for the reaction [Cu(DMF)₄]²⁺ + N ? [Cu(DMF)₃N₃]⁺ + DMF determined by the limiting logarithmic method was found to be 1,25 · 10⁴l · mol^?1. The presence of [Cu(DMF)₄]²⁺ ions in the polymerization systems caused retardation, but [Cu(DMF)₃N₃]⁺ ions produced well defined induction periods. The rate constants at 60°C for the interaction of polystyryl radical towards [Cu(DMF)₄]²⁺ and [Cu(DMF)₃N₃]⁺ ions were calculated to be 6,6 · 10² and 5,74 · 10⁴ l · mol^?1 · s^?1, respectively.     

Trägerkatalysatoren aus C12-allylnickel(II)-komplexen [Ni(C12H19)]X (X = SbF6, O3SCF3) und amorphem aluminiumfluorid in toluol als modell-katalysatoren für das technische katalysatorsystem Ni(octanoat)2/BF3·OEt2/AlEt3 zur 1,4-cis-polymerisation des butadiens

The η³, η², η²-dodeca-2(E), 6(E), 10(Z)-trien-1-yl-nickel(II) complexes [Ni(C₁₂H₁₉)]X (X = SbF₆, O₃SCF₃) were treated in toluene with amorphous aluminium trifluoride (which was prepared from AlEt₃ and BF₃ · OEt₂) in a mole ratio 1 : 10 to 20, forming a highly active catalyst for the 1,4-cis polymerization of butadiene. This catalyst is comparable in its activity and selectivity, and in the molar mass distribution of the polybutadiene, with the technical nickel catalyst Ni(O₂CR)₂/BF₃?OEt₂/AlE₃ developed by Bridgestone Tire Company thirty years ago. The existence of the C₁₂-allynickel(II) cation [Ni(C₁₂H₁₉)]⁺ on the AlF₃ support could be proved by FAB mass spectroscopic measurements. In agreement with our reaction model for the allyl nickel complex catalyzed butadiene polymerization, it is concluded that the technical nickel catalyst in its effective structure can be described as a polybutadienylnickel(II) complex co-ordinated to a polymeric fluoroaluminate anion via a fluoride bridge.     

Catalytic activity of novel agglomerates composed of Fe4S42+ and (MovO)3+ species cross-linked with a bulky difunctional ligand, 2,4,6-trimethyl-1,3-benzenedithiolate

The (Mo^VO)³⁺ -containing [Fe₄S₄]²⁺ agglomerate {(Ph₄P)₂[Fe₄S₄(tmbdt)₂ (Mo^VO)_0,76]}_n (tmbdt = 2,4,6-trimethylbenzene-1,3-dithiolato) was synthesized by reaction of {(Ph₄P)₂[Fe₄S₄(tmbdt)₂]}_n and Mo^VOCl₃(thf)₂. The (Mo^VO)³⁺ cation is coordinated to the sulfur ligands binding the [Fe₄S₄]²⁺ core. The (Mo^VO)³⁺ -containing [Fe₄S₄]²⁺ agglomerate exhibits catalytic activity for the reduction of azobenzene and phenylacetylene to hydrazobenzene and phenylethylene, respectively, by Et₄NBH₄ in N,N-dimethylformamide/alcohol.     

Regulation of Ca channel by intracellular Ca2+ and Mg2+ in frog ventricular cells

The effects of changing the intracellular concentrations of Ca²⁺ or Mg²⁺ ([Ca²⁺]_i, [Mg²⁺]_i) on Ca current (I _Ca) was studied in frog ventricular myocytes using the whole-cell and cell-attached patch clamp techniques. In the physiological range of [Mg²⁺]_i an increase in [Ca²⁺]_i enhancedI _Ca whereas at lower [Mg²⁺]_i I _Ca was suppressed. The increase inI _Ca caused by Ca²⁺ loading was not mediated by phosphorylation since the kinase inhibitors H-8 {N-[2-(methylamino)-ethyl]-5-isoquinolinesulphonamide dihydrochloride}, staurosporine and KN-62 {1-[N,O-bis(5-isoquinoline-sulphonyl)-N-methyl-1-tyrosyl]-4-phenylpiperazine} and a non-hydrolysable adenosine 5-triphosphate analogue ,-methyleneadenosine 5-triphosphate did not prevent the Ca²⁺-inducedI _Ca increase.I _Ca was dramatically increased from 10 ± 6 (n = 4) to 71 ± 7 nA/nF (n = 4) when [Mg²⁺]_i was lowered from 1.0 × 10^–3 to 1.0 × 10^–6 M at a [Ca²⁺]_i of 10^–8 M. The concentration response relation for inhibition of Ca channels by [Mg²⁺]_i is modulated by [Ca²⁺]_i. To account for the experimental results it is postulated that competitive binding of Ca²⁺ or Mg²⁺ to the Ca channel accelerates the transition of the channel from an active to a silent mode. Single-channel recordings support this hypothesis. The regulation may have clinical relevance in cytoprotection during cardiac ischaemia.     

On the 1,4‐cis‐Polymerization of Butadiene with the Highly Active Catalyst Systems Nd(C3H5)2Cl·1.5 THF/Hexaisobutylaluminoxane (HIBAO), Nd(C3H5)Cl2· 2 THF/HIBAO and Nd(C3H5)Cl2·2 THF/Methyl‐aluminoxane (MAO) – Degree of Polymerization,Polydispersity, Kinetics and Catalyst Formation

The allylneodymium chloride complexes Nd(C₃H₅)₂Cl·1.5 THF and Nd(C₃H₅)Cl₂·2 THF can be activated by adding hexaisobutylaluminoxane (HIBAO) or methylaluminoxane (MAO) in a ratio of Al/Nd = 30 for the catalysis of butadiene 1,4‐cis‐polymerization. A turnover frequency (TOF) of about 20 000 mol butadiene/(mol Nd·h) and cis‐selectivity of 95–97% are achieved under standard conditions ([BD]₀ = 2 m, 35°C, toluene). Molecular weight determinations indicate a low polydispersity (M̄_w (LS)/M̄_n (LS) = 1–1.5), the formation of only one polymer chain per neodymium and the linear increase of the degree of polymerization (DP) with the butadiene conversion, as observed for living polymerizations. First indications of chain‐transfer reaction occur only at the highest conversion or degree of polymerization. The rate law r_P = k_P[Nd][C₄H₆]^1.8 is derived for the catalyst system Nd(C₃H₅)₂Cl·1.5 THF/HIBAO and for the system Nd(C₃H₅)Cl₂·2 THF/MAO the rate law r_P = k_P[Nd] [C₄H₆]² with k_P = 3.24 L²/(mol²·s) (at 35°C). Taking into account the Lewis acidity of the alkylaluminoxanes and the characteristic coordination number of 8 for Nd(III) in allyl complexes the formation of an η³‐butenyl‐bis(η⁴‐butadiene)neodymium(III) complex of the composition [Nd(η³‐RC₃H₄)(η⁴‐C₄H₆)₂(X‐{AlOR}_n)₂] is assumed to be a single‐site catalyst for the chain propagation by reaction of the coordinated butadiene via the π‐allyl insertion mechanism and the anti‐cis and syn‐trans correlation to explain the experimental results.     

Contribution of Na+/Ca2+ exchanger to the regulation of myogenic tone in isolated rat small arteries

The contribution of the Na⁺/Ca²⁺ exchanger to the myogenic vascular tone was examined in rat isolated skeletal muscle small arteries (A_SK) with pronounced myogenic tone and mesenteric small arteries (A_MS) with little myogenic tone. Myogenic tone was assessed by the vascular inner diameter at transmural pressures of 40 and 100 mmHg. To depress the Na⁺/Ca²⁺ exchanger, the extracellular Na⁺ concentration ([Na⁺]o) was lowered from 143 to 1.2 mM by substituting choline‐Cl for NaCl. The A_SK developed significant myogenic tone and constricted further in low [Na⁺]o. Nifedipine (1 μM ) reduced both myogenic tone and low [Na⁺]o‐induced contraction. Because the membrane potential of A_SK was not changed by low [Na⁺]o (–35 ± 2 mV at 143 mM [Na⁺]o, ?37 ± 3 mV at 1.2 mM [Na⁺]o), depolarization‐induced Ca²⁺ influx was not a cause of the low [Na⁺]o‐induced contraction. The A_MS did not develop significant myogenic tone. Although low [Na⁺]o also constricted A_MS, the magnitude of constriction was significantly weaker than that in A_SK (17 ± 4 vs. 47 ± 6%, P < 0.01, at 58 mM Na⁺). With Bay K 8644, A_MS developed myogenic tone, and low [Na⁺]o‐induced constriction was significantly increased. In conclusion, Na⁺/Ca²⁺ exchanger may play an important role in regulating myogenic tone, likely via mediating Ca²⁺‐extrusion.     

Oligomerization and copolymerization of γ-butyrolactone — a monomer known as unable to homopolymerize, 1. Copolymerization with ε-caprolactone

Copolymerization of γ-butyrolactone (γBL) with ε-caprolactone (εCL) initiated with aluminium isopropoxide trimer ([Al(OⁱPr)₃]₃, (A₃)) is described. Copolymers with molecular weights (M _n) up to 3 · 10⁴ and containing up to 43 mol-% repeating units derived from γBL are prepared. Their molecular weight is controlled by the concentrations of the consumed comonomers and the starting concentration of initiator {M _n = (86.09 · [γBL]_c + 114.14 · [εCL]_c)/3[Al(OⁱPr)₃] + 60.10}. ¹³C NMR and DSC data are indicative of a pseudoperiodic or random copolymer structure.     

The effect of intracellular pH on cytosolic Ca2+ in HT29 cells

 The influence of intracellular pH (pH_i) on intracellular Ca²⁺ activity ([Ca²⁺]_i) in HT₂₉ cells was examined microspectrofluorometrically. pH_i was changed by replacing phosphate buffer by the diffusible buffers CO₂/HCO₃ ^–or NH₃/NH₄ ⁺ (pH 7.4). CO₂/HCO₃ ^–buffers at 2,5 or 10% acidified pH_i by 0.1, 0.32 and 0.38 pH units, respectively, and increased [Ca²⁺]_i by 8–15 nmol/l. This effect was independent of the extracellular Ca²⁺ activity and the filling state of thapsigargin-sensitive Ca²⁺ stores. Removing the CO₂/HCO₃ ^–buffer alkalinized pH_i by 0.14 (2%), 0.27 (5%), and 0.38 (10%) units and enhanced [Ca²⁺]_i to a peak value of 20, 65, and 143 nmol/l, respectively. Experiments carried out with Ca²⁺-free solution and with thapsigargin showed that the [Ca²⁺]_i transient was due to release from intracellular pools and stimulated Ca²⁺ entry. NH₃/NH₄ ⁺ (20 mmol/l) induced a transient intracellular alkalinization by 0.6 pHunits and increased [Ca²⁺]_i to a peak (Δ [Ca²⁺]_i = 164 nmol/l). The peak [Ca²⁺]_i increase was not influenced by removal of external Ca²⁺, but the decline to basal [Ca²⁺]_i was faster. Neither the phospholipase C inhibitor U73122 nor the inositol 1,4,5-trisphosphate (InsP ₃) antagonist theophylline had any influence on the NH₃/NH₄ ⁺-stimulated [Ca²⁺]_i increase, whereas carbachol-induced [Ca²⁺]_i transients were reduced by more than 80% and 30%, respectively. InsP ₃ measurements showed no change of InsP ₃ during exposure to NH₃/NH₄ ⁺, whereas carbachol enhanced the InsP ₃ concentration, and this effect was abolished by U73122. The pH_i influence on ”capacitative” Ca²⁺ influx was also examined. An acid pH_i attenuated, and an alkaline pH_i enhanced, carbachol- and thapsigargin-induced [Ca²⁺]_i influx. We conclude that: (1) an alkaline pH_i releases Ca²⁺ from InsP ₃-dependent intracellular stores; (2) the store release is InsP ₃ independent and occurs via an as yet unknown mechanism; (3) the store release stimulates capacitative Ca²⁺ influx; (4) the capacitative Ca²⁺ influx activated by InsP ₃ agonists is decreased by acidic and enhanced by alkaline pH_i. The effects of pH_i on [Ca²⁺]_i should be of relevance under many physiological conditions. Received: 17 June 1996 / Received after revision and accepted: 30 August 1996     

Synthesis and characterization of new cardo polyamide-imides containing ether and tricyclo[5.2.1.02, 6]decane groups

A new cardo diimide-dicarboxylic acid, 8,8-bis[4-(4-trimellitimidophenoxy)phenyl]tricyclo-[5.2.1.0^{2, 6}]decane (BTPTD), containing an ether linkage and a tricyclo[5.2.1.0^{2, 6}]decane group was synthesized by condensation reaction of 8,8-bis[4-(4-aminophenoxy)phenyl]tricyclo[5.2.1.0^{2, 6}]decane with trimellitic anhydride. A series of new cardo polyamide-imides were prepared by direct polycondensation of BTPTD with various aromatic diamines in N-methyl-2-pyrrolidinone (NMP) using triphenyl phosphite and pyridine as condensing agents. The polymers were produced with high yield and moderate to high inherent viscosities, i. e., of 0.73–1.36 dL·g^–1. Nearly all polymers are readily soluble in polar aprotic solvents such as NMP, N,N-dimethylacetamide (DMAc), N,N-dimethylformamide, dimethyl sulfoxide as well as less polar solvents such as pyridine, γ-butyrolactone, and tetrahydrofuran. These polymers were solution-cast from DMAc solution into transparent, flexible, and tough films except for polymers 4 a and 4 b . Wide-angle X-ray measurement revealed that all polymers were amorphous. These polyamide-imides have glass transition temperatures between 250–290°C and 5% weight loss temperatures in the range of 445 to 491 and 462 to 490°C in nitrogen and air atmosphere, respectively. The polymer films have a tensile strength range of 77 to 87 MPa, an elongation at break range of 3 to 9%, and a tensile modulus range of 2.2 to 2.8 GPa.     

Ethylene Polymerizations with Alkyl‐, Disilylamino‐ and Cyclopentadienylchromium/MMAO Initiators

The catalytic activities of non‐metallocene type chromium, half‐sandwich type chromium and metallocene type chromium complexes for the polymerization of ethylene were explored using MAO, modified MAO (MMAO) or BuAO in the presence or absence of a silica support. The complexes, Cr[N(SiMe₃)₂]₃ ( 1 ), Cr[CH(SiMe₃)₂]₃ ( 2 ), Cr(CH₂SiMe₃)₄ ( 3 ), and Cr(CH₂CMe₃)₄ ( 4 ), displayed good catalytic activities for the polymerization of ethylene in the presence of excess MAO or MMAO. The activities exceed 10³ g‐PE/Cr‐mol·h to lead to polymers with M_w > 80×10⁴. The molecular weight distributions are rather narrow in cases of complexes 1 and 2 . More efficient catalysis was observed when C₅Me₄‐CH₂CH₂NMe₂CrCl₂ ( 5 ) and C₅H₄CH₂CH₂N(iPr)₂CrCl₂ ( 6 ) were used coupling with MMAO in the presence of a silica support. The M_w increased to 276.2×10⁴ and the M_w/M_w value decreased into 2.2 in the case of complex 5 . The catalysis of (C₅H₅)₂Cr ( 7 ) in the presence of a silica support is very high and the complex ( 7 ) produced high molecular weight polyethylene with narrow molecular weight distribution, while [1,3‐(SiMe₃)₂C₅H₃]₂Cr ( 8 ) shows only a little catalytic activities presumably due to its very weak interaction with a silica support.     

New catalysts for the oxidative polymerization of 2,6-dimethylphenol, 4. The crystal and molecular structure of tetrakis(tetrabutylammonium) hexakis(2,2-dicyano-1,1-ethenedithiolato)octacuprate(i), tetrakis(tetrabutylammonium) hexakis-(1,2-dicyano-1,2-ethenedithiolato)octacuprate(i), and hexakis[copper(i)-n,n-diethylthiocarbamate]

The X-ray determination of the crystal structures shows that the dicyanoethenedithiolatocopper(I) complexes [N(C₄H₉)₄]₄[Cu₈(S₂C?C(CN)₂)₆] ( 1 ) and [N(C₄H₉)₄]₄[Cu₈(S(CN)C?C(CN)S)₆] ( 2 ) contain cube-shaped Cu₈-clusters, whereas the Cu(I)-N,N-(diethyl)thiocarbamate complex ( 3 ) contains a Cu₆-octahedron in its centre. A common feature of the three complexes is the coordination of each copper atom by three donor atoms having suitable empty orbitals for back-donation. This results in flat pyramidal arrangements with the basal plane of the three donor atoms being outside the cluster. All clusters deviate significantly from their ideal symmetry. In complex 2 the Cu₈-cube is tetragonally elongated and the Cu? Cu-distances are in general much larger (3,20 and 3,70 Å, respectively) than in other comparable complexes. Moreover, its anions are fitted into cages of a pseudocubic hydrophobic matrix formed by the tetrabutylammonium cations. It is concluded that is is the bonding within the coordination pyramids of the copper atoms which stabilizes the clusters, whereas the bonding within the clusters seems to be of comparably small importance.     

Increase of intracellular Ca by P2X and P2Y receptor-subtypes in cultured cortical astroglia of the rat

Astrocytes express purinergic receptors that are involved in glial–neuronal cell communication. Experiments were conducted to characterize the expression of functional P2X/P2Y nucleotide receptors in glial cells of mixed cortical cell cultures of the rat. The vast majority of these cells was immunopositive for glial fibrillary acidic protein (GFAP) and was considered therefore astrocyte-like; for the sake of simplicity they were termed “astroglia” throughout. Astroglia expressed predominantly P2X_4,6,7 as well as P2Y_1,2 receptor-subtypes. Less intensive immunostaining was also found for P2X₅ and P2Y_4,6,13,14 receptors. Pressure application of ATP and a range of agonists selective for certain P2X or P2Y receptor-subtypes caused a concentration-dependent increase of intracellular Ca²⁺ ([Ca²⁺]_i). Of the agonists tested, only the P2X_1,3 receptor-selective α,β-methylene ATP was ineffective. Experiments with Ca²⁺-free solution and cyclopiazonic acid, an inhibitor of the endoplasmic Ca²⁺-ATPase, indicated that the [Ca²⁺]_i response to most nucleotides, except for ATP and 2′,3′-O-(benzoyl-4-benzoyl)-ATP, was due primarily to the release of Ca²⁺ from intracellular stores. A Gprotein–mediated release of Ca²⁺ is the typical signaling mechanism of various P2Y receptor-subtypes, whose presence was confirmed also by cross-desensitization experiments and by using selective antagonists. Thus, our results provide direct evidence that astroglia in mixed cortical cell cultures express functional P2Y (P2Y_1,2,6,14 and probably also P2Y₄) receptors. Several unidentified P2X receptors, including P2X₇, may also be present, although they appear to only moderately participate in the regulation of [Ca²⁺]_i. The rise of [Ca²⁺]_i is due in this case to the transmembrane flux of Ca²⁺ via the P2X receptor-channel. In conclusion, P2Y rather than P2X receptor-subtypes are involved in modulating [Ca²⁺]_i of cultured astroglia and thereby may play an important role in cell-to-cell signaling.     

Interrelationship between pH,plasma potassium concentration and ventilation during intense continuous exercise in man

Summary During resting conditions plasma hydrogen ion concentration ([H⁺]_P) is known to influence ventilation , whereas the control of plasma potassium concentration ([K⁺]_P) at rest and of both [K⁺]_P and during exercise are controversial issues. To obtain more information about these variables during muscular work, eight trained men performed two successive intense continuous cycle-ergometer tests, the first (test I) during metabolic acidosis, the second (test II) with an alkalotic pH. No correlation was found between [H⁺]_P and [K⁺]_P or in the direction of change of these variables in test I. Furthermore, no correlation between [H⁺]_P and [K⁺]_P in test I and II was seen. Instead [K⁺]_P and changed in relation to the exercise intensity. We suggest that the results confirm [K⁺]_P as an indicator of muscular stress. In addition, the similar behaviour of relative values of [K⁺]_P ande256-05 changes in test I (r=0.9,m=1.0, wherem is the slope of the regression curve) supports the hypothesis that extracellular potassium controlse256-06 and thereby [H⁺]_P also.     

Modulation of Ca2+ oscillation and apamin-sensitive,Ca2+-activated K+ current in rat gonadotropes

In rat pituitary gonadotropes, gonadotropin-releasing hormone (GnRH) stimulates rhythmic release of Ca²⁺ from stores sensitive to inositol 1,4,5-trisphosphate [Ins(1,4,5)P ₃ ], which in turn induces an oscillatory activation of apamin-sensitive Ca²⁺-activated K⁺ current, I _K(Ca). Since GnRH also activates protein kinase C (PKC), we investigate the action of PKC while simultaneously measuring intracellular Ca²⁺ concentration ([Ca²⁺]_i) and I _K(Ca). Stimulation of PKC by application of phorbol 12-myristate 13-acetate (PMA) did not affect basal [Ca²⁺]_i. However, PMA or phorbol 12,13-dibutyrate (PdBu), but not the inactive 4-phorbol 12,13-didecanoate (4-PDD), reduced the frequency of GnRH-induced [Ca²⁺]_i oscillation and augmented the I _K(Ca) induced by any given level of [Ca²⁺]_i. The slowing of oscillations and the enhancement of I _K(Ca) were mimicked by synthetic diacylglycerol (1,2-dioctanoyl-sn-glycerol) and could be induced during ongoing oscillations that had been initiated irreversibly in cells loaded with guanosine 5-O-(3-thio-triphosphate) (GTP-[S]). In contrast, when oscillations were initiated by loading cells with Ins(1,4,5)P ₃, phorbol esters enhanced I _K(Ca) without affecting the frequency of oscillation. The protein kinase inhibitor, staurosporine, reduced I _K(Ca) without affecting [Ca²⁺]_i and partially reversed the phorbol-ester-induced slowing of oscillation. Therefore, activation of PKC has two rapid effects on gonadotropes. It slows [Ca²⁺]_i oscillations probably by actions on phospholipase C, and it enhances I _K(Ca) probably by a direct action on the channels.     

Antidiuretic hormone acts via V1 receptors on intracellular calcium in the isolated perfused rabbit cortical thick ascending limb

The effect of antidiuretic hormone ([Arg]vasopressin, ADH) on intracellular calcium activity [Ca²⁺]_i of isolated perfused rabbit cortical thick ascending limb (cTAL) segments was investigated with the calcium fluorescent dye fura-2. The fluorescence emission ratio at 500–530 nm (R) was monitored as a measure of [Ca²⁺]_i after excitation at 335 nm and 380 nm. In addition the transepithelial potential difference (PD _te) and transepithelial resistance (R _te) of the tubule were measured simultaneously. After addition of ADH (1–4 nmol/l) to the basolateral side of the cTAL R increased rapidly, but transiently, from 0.84±0.05 to 1.36±0.08 (n = 46). Subsequently, within 7–12 min R fell to control values even in the continued presence of ADH. The increase in R evoked by the ADH application corresponded to a rise of [Ca²⁺]_i from a basal level of 155±23 nmol/l [Ca²⁺]_i up to 429±53 nmol/l [Ca²⁺]_i at the peak of the transient, as estimated by intra- or extracellular calibration procedures. The electrical parameters (PD _te and R _te) of the tubules were not changed by ADH. The ADH-induced Ca²⁺ transient was dependent on the presence of Ca²⁺ on the basolateral side, whereas luminal Ca²⁺ had no effect. d(CH₂)₅[Tyr(Me)²]²,Arg⁸vasopressin, a V₁ antagonist (Manning compound, 10 nmol/l), blocked the ADH effect on [Ca²⁺]_i completely (n = 5). The V₂ agonist 1-desamino-[d-Arg⁸]vasopressin (10 nmol/l, n=4), and the cAMP analogues, dibutyryl-cAMP (400 mol/l, n = 4), 8-(4-chlorophenylthio)-cAMP (100 mol/l, n = 1) or 8-bromo-cAMP (200 mol/1, n = 4) had no influence on [Ca²⁺]_i. The ADH-induced [Ca²⁺]_i increase was not sensitive to the calcium-channel blockers nifedipine and verapamil (100 mol/l, n = 4). We conclude that ADH acts via V₁ receptors to increase cytosolic calcium activity transiently in rabbit cortical thick ascending limb segments, possibly by an initial Ca²⁺ release from intracellular stores and by further Ca²⁺ influx through Ca²⁺ channels in the basolateral membrane. These channels are insensitive to L-type Ca²⁺ channel blockers, e.g. nifedipine and verapamil.Supported by DFG GR 480/10