Hydrogen peroxide-induced Ca responses in CNS pericytes

Objective: The aims of the present study were to elucidate the interaction of reactive oxygen species (ROS) and Ca²⁺ response in central nervous system (CNS) pericytes. Methods: The intracellular Ca²⁺ concentration was measured using fluorescent Ca²⁺ indicator, fura-2, in cultured CNS pericytes. Results: Hydrogen peroxide evoked a dose-dependent increase in cytosolic Ca²⁺, which was completely inhibited by catalase. Removal of external Ca²⁺ or addition of nicardipine (1 μM) during application of hydrogen peroxide did not affect Ca²⁺ response. Incubation of the cells in Ca²⁺ free solution did not abolish but slightly reduced Ca²⁺ response by hydrogen peroxide. Ca²⁺ response to hydrogen peroxide was not altered by the depletion of intracellular Ca²⁺ by thapsigargin (1 μM). Pretreatment of the cells with tyrosine kinase inhibitor genistein (100 μM) or tyrphostin A47 (30 μM) significantly reduced Ca²⁺ increase by hydrogen peroxide. Conclusions: These results indicate that hydrogen peroxide evokes Ca²⁺ increase predominantly by release from intracellular Ca²⁺ store, which may be regulated by tyrosine kinases.     

Caffeine induces periodic oscillations of Ca2+-activated K+ current in pulmonary arterial smooth muscle cells

The periodic oscillations of outward currents were studied in smooth muscle cells of the rabbit pulmonary artery. The combined stimuli of superfusion with 1 mM caffeine and depolarization of the membrane potential to 0 mV evoked periodic oscillations of outward currents with fairly uniform amplitudes and intervals. The oscillating outward currents induced by caffeine were dependent on intracellular Ca²⁺ concentration ([Ca²⁺]_i) and had a reversal potential near to the equilibrium potential for K⁺. So the oscillating outward currents are carried by K⁺ through Ca²⁺-dependent K⁺ channels (I _K(Ca)), and may reflect the oscillations of [Ca²⁺]_i. The oscillating outward currents were abolished, or their frequency reduced, by lowering external [Ca²⁺], Ca²⁺ channel blockers, or by 1 M ryanodine, indicating that: (1) there is a continuous influx of Ca²⁺ through the plasma membrane at a holding potential of 0 mV; (2) the periodic transient increases of [Ca²⁺]_i are ascribed to the rhythmic release of Ca²⁺ from ryanodinesensitive intracellular store by the mechanism of Ca²⁺-induced Ca²⁺ release (CICR). On the basis of the above results, we simulated the oscillation of [Ca²⁺]_i induced by caffeine, which is known to lower the threshold of CICR. The patterns of peak amplitude histograms of spontaneous transient outward currents (STOC) in the oscillating cells were different from those in non-oscillating cells. The amplitudes of STOC in the latter were more variable than those in the former. The oscillating outward currents were modulated by 1 M forskolin and 1 M sodium nitroprusside, but STOC were little affected. The above differences between STOC and oscillating outward currents suggest that the two currents are activated by the Ca²⁺ originating from different intracellular Ca²⁺ stores which are functionally heterogeneous.     

Changes in cyclic nucleotides during the calcium paradox in the isolated rat heart

Reperfusion of hearts with a Ca²⁺-containing medium after a perfusion period in Ca²⁺-free medium results in irreversible cell damage (calcium paradox). In this investigation we have studied coronary flow and cyclic AMP and cyclic GMP levels after several periods of Ca²⁺-free perfusion in isolated rat hearts. We also investigated the effects of papaverine (Pap), noradrenaline (NA), acetylcholine (ACh) and absence of inorganic phosphate during Ca²⁺-free perfusion on coronary flow (CF) and cyclic nucleotide levels. Inability of the heart to recover contractile activity with development of contracture during the reperfusion period was accepted as indicative of the calcium paradox. Ca²⁺-free perfusion alone and NA and absence of inorganic phosphate during the Ca²⁺-free perfusion period increased CF, whereas Pap and ACh decreased it. However, only Ca²⁺-free perfusion and NA elevated cyclic AMP. On the other hand, Pap and ACh increased cyclic GMP (with a transient rise of cyclic AMP in Pap infusion), and absence of inorganic phosphate decreased both cyclic AMP and cyclic GMP. Pap, ACh and absence of phosphate prevented the calcium paradox. Our study suggests that increased cyclic AMP during the Ca²⁺-free perfusion may contribute, with the other factors, to the occurrence of the calcium paradox.     

Characterization of a whole-cell Ca2+-blockable monovalent cation current in isolated ectodermal cells of chick embryo

The presence of a Ca²⁺-blockable monovalent cation current is demonstrated in isolated ectodermal cells of the chick embryo using the whole-cell patch-clamp method. In the absence of any stimulation, the whole-cell current is time independent and rectifies outwardly at membrane potentials higher than +40 mV The outward current is neither carried by Cl^– channels nor by K⁺ channels. Application of a Ca²⁺-free solution containing 1 mmol/l ethylenediaminetetraacetic acid (EDTA) elicits a large inward current and increases the outward current. The inward current can be carried by extracellular Li⁺, Na⁺, K⁺ and Cs⁺, but notN-methyl-d-glucamine. The Ca²⁺-blockable monovalent cation channel discriminates very poorly among these cations. The estimated number of channels per cell is around 2000. Extracellular protons block the inward Na⁺ current in the absence of extracellular Ca²⁺. The apparent negative logarithm of the dissociation constant for proton (pK _H) at –100 mV is 5.8. Among 12 potential channel modulators, including verapamil and nifedipine, only quinine decreases the current. Quinine blocks this current with a dissociation constant,K _d, equal to 0.18 mmol/l, independent of the membrane potential. This study demonstrates the presence of a whole-cell Ca²⁺-blockade monovalent cation current in dissociated chick ectodermal cells with permeation properties similar to those observed at the single-channel level. Contrary to studies made of other tissues, we did not observe any blocking effect of verapamil and nifedipine on the Ca²⁺-blockable monovalent cation current.     

Ampullary electroreceptors in catfish (Teleostei): temperature dependence of stimulus transduction

The response properties of ampullary electroreceptors have been studied in the catfish Ictalurus nebulosus at skin temperatures between 5 and 35 °C. A unimodal relationship between spontaneous activity and temperature was obtained. Mean (±SEM) peak discharge rate was 57.3 ±1.8 impulses s^–1 at 25 ° C; the receptors were active at 5 °C (15.0 impulses s^–1) and at 35 °C (31.5 impulses s^–1). There were no dynamic responses to temperature changes in either the warming or cooling direction. The shape of the frequency characteristic depended on temperature: the peak of the gain curve shifted to low frequencies at low temperatures. There was a concomitant change of the phase characteristic: the intersection at zero degree phase angle shifted to higher frequencies with an increase of temperature, thus increasing the lead at lower frequencies and decreasing the lag at higher frequencies. Latency after combined excitatory and inhibitory impulse stimulation was temperature dependent, ranging from 16.4 ms (5 °C) to 5.6 ms (35 °C). Application of the specific calcium channel blocker menthol (0.2 mM) suppressed spontaneous activity, the effect becoming more prominent at higher temperatures. Sensitivity to sinusoidal electrical stimulation was also impaired, but to a lesser degree and mainly at lower temperatures. We conclude that the filter properties of the receptor organ can be modelled by a band-pass filter in series with a latency, both of which are temperature dependent. These filter properties might be partially based on the activation kinetics of the tranduction channels.     

A method for studying inhibitory activity in whole urine

Summary A method has been developed for inducing and quantifying calcium oxalate crystallisation in whole human urine. The propensity of a given urine to induce crystal formation was described in two ways: 1) its ability to resist spontaneous nucleation of calcium oxalate crystals was assessed by titrating 20 mls of the urine with increasing quantities of sodium oxalate (0–150 mol) to determine its practical metastable limit. This limit was inversely related to the endogenous calcium concentration. 2) its capacity to inhibit crystal growth was quantified by determining the rate of growth of calcium oxalate crystals precipitated in response to a fixed oxalate load (30 mol) above its metastable limit. The crystals produced were predominantly calcium oxalate dihydrate and were morphologically identical to those occurring naturally in urine. Citrate had no effect on the metastable limits of 3 urines examined, but markedly inhibited crystal growth. Pyrophosphate had a similar effect on crystal growth, and in addition, raised the metastable limit of one of the urine samples.     

Nifedipine and alpha1-adrenergic blockade in Raynaud's phenomenon

The efficacy of nifedipine and prazosin in the treatment ofRaynaud's phenomenon was assessed in a prospective double-blindrandomized cross-over trial in 15 patients. Each patient receivedone week of nifedipine 20 mg TID, one week of prazosin 1 mgTID, and 2 weeks of placebo. Nifedipine was shown to be effectivein reducing both the frequency and the severity of Raynaud'sphenomenon, whereas prazosin was ineffective. Before initiationof therapy in the 15 patients, pressor responses to the intravenousalpha₁-agonist phenylephrine were assessed in the basal state,30 min after 20 mg oral nifedipine, and 30 min after 1 mg oralprazosin; the shift to the right of the log dose-vasopressorresponse curves to phenylephrine was similar with nifedipineand prazosin.     

The effects of verapamil on cerebrospinal fluid pressure in surgical patients

The effects of verapamil upon cerebrospinal fluid pressure (CSFP) were studied in twenty surgical patients without intracranial pathology who were divided into two groups of ten patients each: verapamil 0.075mg·kg^–1 was given in group 1 and 0.15mg·kg^–1 was given in group 2. A spinal needle was inserted into the subarachnoid space to permit continuous measurement of CSFP. Intravenous verapamil as a bolus produced a statistically significant increase in CSFP: from 6.0 ± 3.5 (mean ± SD) to 10.5 ± 4.3mmHg in group 1 (P < 0.01), and from 6.2 ± 3.1 to 12.6 ± 3.8mmHg in group 2 (P < 0.01). CSFP after verapamil attained its maximum in 0.5–1.5min, then gradually returned to control levels. Changes in CSFP were always associated with statistically significant decreases in arterial blood pressure and cerebral perfusion pressure, while the heart rate showed variable changes. It is concluded that a clinical dose of verapamil showed variable changes. It is concluded that a clinical dose of verapamil (0.075–0.15mg·kg^–1) has no neurological side effects in patients without intracranial hypertension. However, it must be emphasized that verapamil may increase CSFP to undesirable levels and should be avoided in patients with compromised intracranial compliance.(Nishikawa T, Namiki A: The effects of verapamil on cerebrospinal fluid pressure in surgical patients. J Anesth 1: 132–136, 1987)     

Pharmacological modification of tremor and enhanced acoustic startle by chlordecone and p,p′-DDT

Pretreatment of rats with phenoxybenzamine (5 mg/kg; SC), an alpha adrenergic antagonist, decreased the peak tremor power and startle magnitude of rats subsequently given DDT (75 mg/kg; PO) or chlordecone (60 mg/kg; IP), without having a significant effect on control animals. Pretreatment with an intracerebroventricular injection of calcium (3.75 M in 5 l NaCl) decreased the peak tremor power due to subsequently administered DDT, while increasing the tremor response in rats later dosed with chlordecone. The effects of phenoxybenzamine are postulated to be due to a blockade of an excitatory influence of the adrenergic system. Calcium may decrease DDT-induced tremor by acting as a neuronal stabilizer. Potentiation of the tremorigenic effect of chlordecone by calcium may be due to increased levels of intracellular calcium, resulting in augmented release of neurotransmitters in chlordecone-exposed animals.     

Crystalluria in marathon runners

Summary Epidemiological evidence suggests that marathon runners have a higher incidence of renal stone formation than occurs in the general population. Since crystalluria and stone disease are thought to be related, we subjected urine samples from a group of marathon runners to particle counting and sizing in a Coulter Counter equipped with a population accessory unit. The volume-size distribution curves so obtained were bimodal with one peak occurring in the 2–5 m diameter range and a second in the 15–32 m diameter range—a pattern that is remarkably similar to the distributions reported for recurrent idiopathic stone formers and distinctly different to those recorded for control subjects. Analyses by scanning electron microscopy and X-ray powder diffraction revealed other features which are regarded as typical of stone formers' crystalluria. These physicochemical data indicate that marathon runners may be at increased risk of urinary stone formation.