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I Parker

Publications and source records attributed to I Parker.

At least 37 records · Page 2Linked to original sources
PubMed1998

Hemispheric asymmetry of macroscopic and elementary calcium signals mediated by InsP3 in Xenopus oocytes.

1. The mechanisms underlying hemispheric asymmetry of the inositol 1, 4,5-trisphosphate (InsP3)-calcium signalling pathway in Xenopus oocytes were examined by fluorescence imaging of calcium signals and recording calcium-activated Cl- currents (ICl,Ca) evoked by intracellular calcium injections and photorelease of InsP3. 2. The maximal ICl,Ca evoked by strong photorelease of InsP3 was 8 times greater in the animal than the vegetal hemisphere, but the average threshold amounts of InsP3 required to evoke detectable currents were similar in each hemisphere. 3. Currents evoked by injections of calcium were about 2.5 times greater near the animal pole than near the vegetal pole, whereas fluorescence signals evoked by injections were similar in each hemisphere. 4. Calcium waves were evoked by photolysis flashes of similar strengths in both hemispheres of albino oocytes, but peak calcium levels evoked by supramaximal stimuli were 70 % greater in the animal hemisphere. 5. Elementary calcium release events (puffs) in the animal hemisphere had amplitudes about double that in the vegetal hemisphere, and more often involved coupled release from adjacent sites. Calcium release sites were more closely packed in the animal hemisphere, with a mean spacing of about 1.5 micro m compared with 2.25 micro m in the vegetal hemisphere. 6. The larger amplitude of currents mediated by InsP3 in the animal hemisphere, therefore, involves an increased flux of calcium at individual release units, a more dense packing of release units and a higher density of Cl- channels.

Animals↗
PubMed1998

A continuum of InsP3-mediated elementary Ca2+ signalling events in Xenopus oocytes.

1. The elementary release events underlying inositol 1,4, 5-trisphosphate (InsP3)-mediated calcium signalling were investigated in Xenopus oocytes by means of high-resolution confocal linescan imaging together with flash photolysis of caged InsP3. 2. Weak photolysis flashes evoked localized, transient calcium signals that arose at specific sites following random latencies of up to several seconds. The duration, spatial spread and amplitude of these elementary events varied widely. Event durations (at half-maximal amplitude) were distributed exponentially between about 100 and 600 ms. Fluorescence magnitudes (F/F0 of Oregon Green 488 BAPTA-1) showed a skewed distribution with a peak at about 1.5 and a tail extending as high as 3.5. 3. Individual release sites exhibited both small events (blips) and large events (puffs). The spatiotemporal distribution of calcium signals during puffs was consistent with calcium diffusion from a point source (< a few hundred nanometres), rather than with propagation of a microscopic calcium wave. 4. Estimates of the calcium flux associated with individual events were made by integrating fluorescence profiles along the scan line in three dimensions to derive the 'signal mass' at each time point. The smallest resolved events corresponded to liberation of < 2 x 10-20 mol Ca2+, and large events to about 2 x 10-18 mol Ca2+. The rise of signal mass was more prolonged than that of the fluorescence intensity, suggesting that calcium liberation persists even while the fluorescence begins to decline. Rates of rise of signal mass corresponded to Ca2+ currents of 0.4-2.5 pA. 5. Measurements of signal mass from different events showed a continuous, exponential distribution, arising through variability in magnitude and duration of calcium flux. 6. We conclude that localized calcium transients in the oocyte represent a continuum of events involving widely varying amounts of calcium liberation, rather than falling into separate populations of 'fundamental' and 'elementary' events (blips and puffs) involving, respectively, single and multiple InsP3 receptor channels. This variability probably arises through stochastic variation in both the number of channels recruited and the duration of channel opening.

Animals↗
PubMed1998

Activation and co-ordination of InsP3-mediated elementary Ca2+ events during global Ca2+ signals in Xenopus oocytes.

1. The activation of elementary calcium release events ('puffs') and their co-ordination to generate calcium waves was studied in Xenopus oocytes by confocal linescan imaging together with photorelease of inositol 1,4,5-trisphosphate (InsP3) from a caged precursor. 2. Weak photolysis flashes evoked no responses or isolated calcium puffs, whereas flashes of increasing strength evoked more frequent puffs, often occurring in flurries as abortive waves, and then a near-simultaneous calcium liberation originating at multiple sites. The numbers of sites activated increased initially as about the fourth power of photoreleased [InsP3]. 3. Following repeated, identical photolysis flashes, puffs arose after stochastically varying latencies of a few hundred milliseconds to several seconds. The cumulative number of events initially increased as about the third power of time. No rise in free [Ca2+] was detected preceding the puffs, suggesting that this co-operativity arises through binding of multiple InsP3 molecules, rather than through calcium feedback. 4. The mean latency to onset of calcium liberation shortened as about the square of the flash strength, and the dispersion in latencies between events reduced correspondingly. 5. Weak stimuli often evoked coupled puffs involving adjacent sites, and stronger flashes evoked saltatory calcium waves, propagating with non-constant velocity. During waves, [Ca2+] rose slowly between puff sites, but more abruptly at active sites following an initial diffusive rise in calcium. 6. Initial rates of rise of local [Ca2+] at release sites were similar during puffs and release induced by much (> 10-fold) greater [InsP3]. In contrast, macroscopic calcium measurements averaged over the scan line showed a graded dependence of rate of calcium liberation upon [InsP3], due to recruitment of additional sites and decreasing dispersion in activation latencies. 7. We conclude that the initiation of calcium liberation depends co-operatively upon [InsP3] whereas the subsequent regenerative increase in calcium flux depends upon local calcium feedback and is largely independent of [InsP3]. Wave propagation is consistent with the diffusive spread of calcium evoking regenerative liberation at heterogeneous discrete sites, the sensitivity of which is primed by InsP3.

Animals↗
PubMed1997

Variability in frequency and characteristics of Ca2+ sparks at different release sites in rat ventricular myocytes.

1. High spatial resolution confocal imaging was used to investigate spontaneous calcium release events (Ca2+ sparks) in isolated rat cardiac myocytes loaded with the fluorescent calcium indicator fluo-3. 2. Frequencies of sparks at different release sites varied widely, with a few sites showing sustained activities as great as 50 times the average. Sites with frequent sparks showed more rapid recovery of activity following Ca2+ waves and locally elevated basal [Ca2+]. 3. In addition to transient sparks, some sites showed bursts of rapid flurries of spark-like events, or apparently sustained Ca2+ liberation. Bursts remained localized to individual z-lines, but adjacent sites on the same z-line could be 'driven' by a bursting site to generate similar activity. 4. Individual sites showed long-term (tens of seconds) changes in 'modes' of activity, with abrupt transitions in frequencies of sparking, and between transient sparks and sustained bursts. These transitions were not associated with changes in the amplitude of the sparks. 5. We conclude that spontaneous sparks are not stereotyped events generated with uniform probability at all sites. Instead, the Ca2+ release event in variable, and some sites have a high probability of spark generation. Both factors show long-term changes at individual sites, raising the possibility that properties of fundamental Ca2+ release units may be subject to modulation.

Aniline Compounds↗
PubMed1997

A high-resolution, confocal laser-scanning microscope and flash photolysis system for physiological studies.

We describe the construction of a high-resolution confocal laser-scanning microscope, and illustrate its use for studying elementary Ca2+ signalling events in cells. An avalanche photodiode module and simple optical path provide a high efficiency system for detection of fluorescence signals, allowing use of a small confocal aperture giving near diffraction-limited spatial resolution (< 300 nm lateral and < 400 nm axial). When operated in line-scan mode, the maximum temporal resolution is 1 ms, and the associated computer software allows complete flexibility to record line-scans continuously for long (minutes) periods or to obtain any desired pixel resolution in x-y scans. An independent UV irradiation system permits simultaneous photolysis of caged compounds over either a uniform, wide field (arc lamp source) or at a tightly focussed spot (frequency-tripled Nd:YAG laser). The microscope thus provides a versatile tool for optical studies of dynamic cellular processes, as well as excellent resolution for morphological studies. The confocal scanner can be added to virtually any inverted microscope for a component cost that is only a small fraction of that of comparable commercial instruments, yet offers better performance and greater versatility.

Animals↗
PubMed1997

The effect of azodicarbonamide concentrations on ethyl carbamate concentrations in bread and toast.

A series of baking experiments have been undertaken in order to test the proposition that the use of the flour improver azodicarbonamide influences ethyl carbamate concentrations in baked bread. Samples were prepared in a laboratory and contained 0, 20 and 45 mg azodicarbonamide/kg; 20 mg/kg reflecting normal commercial usage and 45 mg/kg the UK statutory limit. Samples incorporating 0 and 20 mg/kg of the additive were also prepared in a commercial bakery. Toast made from these breads was examined since it is known that toasting can lead to increased ethyl carbamate concentrations. Statistical analysis of the data indicated that, at 45 mg/kg, azodicarbonamide led to significant increases in ethyl carbamate concentrations in both bread and the toasts made from it. At 20 mg/kg some small increases in ethyl carbamate were seen for bread and this approached statistical significance for those samples made in the commercial plant. When these breads were toasted an increase in ethyl carbamate was observed but this was not attributable to the use of azodicarbonamide.

Azo Compounds↗
PubMed1997

The contribution of azodicarbonamide to ethyl carbamate formation in bread and beer.

Data on ethyl carbamate concentrations in beers purchased and analysed between 1988 and 1990 are presented. The concentrations in draught beers were uniformly below the detection limit of 1 microgram/l. Canned beers contained rather more ethyl carbamate (up to 2.5 micrograms/l) which is considered to be due to their longer shelf-life and higher alcohol content (in some cases). Bottled beers contained even higher amounts of ethyl carbamate (up to 14.7 micrograms/l) and this was considered to be due to the use of azodicarbonamide as a blowing agent in the beer bottle cap liners. It is understood that modifications to the liner have led to reduced concentrations in bottled beers produced more recently. A survey of bread samples and related cereal products such as rusks, French toasts and pitta bread indicated typical ethyl carbamate concentrations between < 0.4 and 4.5 micrograms/kg. Toasting bread led to increases of between three- and eight-fold in ethyl carbamate concentrations ranging from 3.5 to 33.8 micrograms/kg on a wet weight basis. Analysis of the data indicated that commercial bread samples which indicated the use of azodicarbonamide as a flour improver showed statistically significant increases in ethyl carbamate concentrations. The mean increase for treated bread over untreated bread was 66%. When these breads were toasted, the mean increase for treated toast over untreated toast was 56%.

Azo Compounds↗
PubMed1996

Ca2+ sparks involving multiple Ca2+ release sites along Z-lines in rat heart cells.

1. High spatial resolution confocal imaging was used to investigate the fundamental nature of "Ca2+ sparks' in rat cardiac myocytes loaded with the fluorescent calcium indicator, fluo-3. 2. The sites at which calcium sparks occurred (Ca2+ release sites) were packed closely and irregularly in transverse planes along Z-lines (mean spacing between sites of 0.76 microns). In contrast, sites were spaced more regularly in the longitudinal direction, at intervals of 1.8 microns (i.e. the sarcomere length). 3. Diffusion of released Ca2+ was slower transversely (apparent diffusion coefficient, D, 7.9 microns 2 s-1) than longitudinally (D, 17.1 microns 2 s-1). 4. Frequently, discrete sites several hundred nanometres apart transversely activated in near synchrony. The probability of transverse synchronous activity fell to low levels (< 20%) at sites separated by more than 1.0 micron. Synchronous activation was not observed between sites on different Z-lines (i.e. separated longitudinally by 1.8 microns). 5. High temporal resolution confocal microscopy (stationary spot) revealed Ca2+ sparks with "stepped' rises, consistent with multiple sites of origin. 6. We conclude that the Ca2+ spark as originally described is usually not an "elementary' event, in the sense of being indivisible, but is often comprised of yet smaller, triggered units of Ca2+ release.

Aniline Compounds↗
PubMed1996

Ca2+ transients associated with openings of inositol trisphosphate-gated channels in Xenopus oocytes.

1. The mechanisms underlying inositol 1,4,5-trisphosphate (InsP3)-induced Ca2+ liberation were studied in Xenopus oocytes by using scanning and stationary-point confocal fluorescence microscopy to record Ca2+ signals evoked by photorelease of InsP3 from a caged precursor. 2. Fluorescence measurements from confocal images showed that increasing [InsP3] evoked three distinct modes of Ca2+ liberation: a diffuse 'pacemaker' signal, localized transient puffs, and propagating waves. Peak free Ca2+ concentrations during waves and puffs (respectively, 2-5 microM and 100-200 nM) varied only slightly with [InsP3], whereas the pacemaker amplitude varied over a wider range (at least 1-30 nM Ca2+). 3. The improved resolution provided by confocal point recording revealed discontinuous Ca2+ 'blips' during pacemaker release. These events were resolved only at particular locations and had time courses similar to the puffs (rise, approximately 50 ms; decay, a few hundred milliseconds) but with amplitudes one-fifth or less of puff amplitudes. 4. We conclude that blips may arise through opening of single InsP3-gated channels, whereas puffs reflect the concerted opening of several clustered channels due to local regenerative feedback by Ca2+.

Animals↗
PubMed1996

Fast kinetics of calcium liberation induced in Xenopus oocytes by photoreleased inositol trisphosphate.

Inositol 1,4,5-trisphosphate (InsP3) acts on intracellular receptors to cause liberation of Ca2+ ions into the cytosol as repetitive spikes and propagating waves. We studied the processes underlying this regenerative release of Ca2+ by monitoring with high resolution the kinetics of Ca2+ flux evoked in Xenopus oocytes by flash photolysis of caged InsP3. Confocal microfluorimetry was used to monitor intracellular free [Ca2+] from femtoliter volumes within the cell, and the underlying Ca2+ flux was then derived from the rate of increase of the fluorescence signals. A threshold amount of InsP3 had to be photoreleased to evoke any appreciable Ca2+ signal, and the amount of liberated Ca2+ then increased only approximately fourfold with maximal stimulation, whereas the peak rate of increase of Ca2+ varied over a range of nearly 20-fold, reaching a maximum of approximately 150 microMs-1. Ca2+ flux increased as a first-order function of [InsP3]. Indicating a lack of cooperativity in channel opening, and was half-maximal with stimuli approximately 10 times threshold. After a brief photolysis flash, Ca2+ efflux began after a quiescent latent period that shortened from several hundred milliseconds with near-threshold stimuli to 25 ms with maximal flashes. This delay could not be explained by an initial "foot" of Ca2+ increasing toward a threshold at which regenerative release was triggered, and the onset of release seemed too abrupt to be accounted for by multiple sequential steps involved in channel opening. Ca2+ efflux increased to a maximum after the latent period in a time that reduced from > 100 ms to approximately 8 ms with increasing [InsP3] and subsequently declined along a two-exponential time course: a rapid fall with a time constant shortening from > 100 ms to approximately 25 ms with increasing [InsP3], followed by a much smaller fail persisting for several seconds. The results are discussed in terms of a model in which InsP3 receptors must undergo a slow transition after binding InsP3 before they can be activated by cytosolic Ca2+ acting as a co-agonist. Positive feedback by liberated Ca2+ ions then leads to a rapid increase in efflux to a maximal rate set by the proportion of receptors binding InsP3. Subsequently, Ca2+ efflux terminates because of a slower inhibitory action of cytosolic Ca2+ on gating of InsP3 receptor-channels.

Animals↗
PubMed1996

Elementary events of InsP3-induced Ca2+ liberation in Xenopus oocytes: hot spots, puffs and blips.

Liberation of sequestered Ca2+ ions in Xenopus oocytes by the second messenger inositol 1,4,5-trisphosphate (InP3) occurs from functionally discrete sites, which are spaced at intervals of several microns and probably represent clusterings of InsP3 receptor/channels (InsP3R) in the endoplasmic reticulum. As well as requiring InsP3, opening of release channels is regulated by dual positive and negative feedback by cytosolic Ca2+, leading to regenerative Ca2+ transients. Because the sensitivity of this process is determined by [InsP3], the ability of Ca2+ ions diffusing from one location to activate increasingly distant InsP3R is enhanced by increasing [InsP3]. Together with the spatial distribution of receptors, this results in generation of a hierarchy of Ca2+ release events, which may involve individual InsP3R (Ca2+ 'blips'), concerted activation of several receptors within a single release site (Ca2+ 'puffs'), and recruitment of successive sites by Ca2+ diffusing over micron distances to produce propagating Ca2+ waves. Thus, Ca2+ signalling in the oocyte is organized as at least two sizes of elemental 'building blocks'; highly localized Ca2+ transients that arise autonomously and stochastically from discrete sites at low [InsP3], but which become coordinated at higher [InsP3] to produce global Ca2+ responses.

Animals↗
PubMed1995

Quantal puffs of intracellular Ca2+ evoked by inositol trisphosphate in Xenopus oocytes.

1. Ca2+ liberation induced in Xenopus oocytes by a poorly metabolized derivative of inositol 1,4,5-trisphosphate (3-deoxy-3-fluoro-D-myo-inositol 1,4,5-trisphosphate; 3-F-InsP3) was visualized using a video-rate confocal microscope to image fluorescence signals reported by the indicator dye calcium green-1. 2. Low (10-30 nM) intracellular concentrations of 3-F-InsP3 evoked Ca2+ release as localized transient 'puffs'. Progressively higher concentrations (30-60 nM) gave rise to abortive Ca2+ waves triggered by puffs, and then (> 60 nM) to a sustained elevation of Ca2+ followed by the appearance of propagating Ca2+ waves. At concentrations up to that giving waves, the frequency of puffs increased as about the third power of [InsP3], whereas their amplitudes increased only slightly. 3. The rise of cytosolic Ca2+ during a puff began abruptly, and peaked within about 50 ms. The peak free Ca2+ level was about 180 nM, and the total amount of Ca2+ liberated was several attomoles (10(-18) mol), too much to be accounted for by opening of a single InsP3-gated channel. The subsequent decline of Ca2+ occurred over a few hundred milliseconds, determined largely by diffusion of Ca2+ away from the release site, rather than by resequestration. Lateral spread of Ca2+ was restricted to a few micrometres, consistent with an effective diffusion coefficient for Ca2+ ions of about 27 microns2 s-1. 4. The peak amplitudes of puffs recorded at a given site were distributed in a roughly Gaussian manner, and a small proportion of sites consistently gave puffs much larger than the main population. Intervals between successive puffs at a single site were exponentially distributed, except for a progressive fall-off in puffs seen at intervals shorter than about 10 s. Thus, triggering of puffs appeared to be stochastically determined after recovery from a refractory period. 5. There was little correlation between the occurrence of puffs at sites more than a few micrometres apart, indicating that puff sites can function autonomously, but closely (ca 2 microns) adjacent sites showed highly correlated behaviour. 6. Puffs arose from sites-present at a density of about 1 per 30 microns2 in the animal hemisphere, located within a narrow band about 5-7 microns below the plasma membrane. 7. We conclude that Ca2+ puffs represent a 'quantal' unit of InsP3-evoked Ca2+ liberation, which may arise because local regenerative feedback by cytosolic Ca2+ ions causes the concerted opening of several closely clustered InsP3 receptor channels.

Animals↗
PubMed1995

Calcium puffs in Xenopus oocytes.

The second messenger inositol 1,4,5-trisphosphate (InsP3) functions in large part by liberating calcium ions from intracellular stores. This release process is highly non-linear and shows a regenerative characteristic that allows production of all-or-none calcium spikes which propagate as waves. However, at low concentrations of InsP3 an additional mode of calcium liberation is seen in Xenopus oocytes, transient 'puffs' of cytosolic calcium that last for a few hundred milliseconds and are restricted to within a few micrometres. Puffs are generally of similar size and the amount of calcium released (about 3 x 10(-18) mol) suggests that they arise through the concerted opening of several InsP3-gated calcium release channels. Puff sites are present at a density of about one per 30 microns 2 in the animal hemisphere of the oocyte. Each site functions autonomously, producing puffs at largely random intervals. We conclude that calcium puffs represent 'quantal' units of InsP3-evoked calcium liberation, which may result from local regenerative feedback by cytosolic calcium ions at functionally discrete release sites.

Animals↗
PubMed1994

Role of cytosolic Ca2+ in inhibition of InsP3-evoked Ca2+ release in Xenopus oocytes.

1. Calcium liberation induced in Xenopus oocytes by flash photorelease of inositol 1,4,5-trisphosphate (InsP3) from a caged precursor was monitored by confocal microfluorimetry. The object was to determine whether inhibition of Ca2+ release seen with paired flashes arose as a direct consequence of elevated cytosolic free [Ca2+]. 2. Responses evoked by just-suprathreshold test flashes were not inhibited by subthreshold conditioning flashes, but were strongly suppressed when conditioning flashes were raised above threshold. 3. Inhibition at first increased progressively as the inter-flash interval was lengthened to about 2 s and thereafter declined, with a half-recovery at about 4 s. 4. Intracellular injections of Ca2+ caused relatively slight inhibition of InsP3-evoked signals, even when cytosolic free [Ca2+] was elevated to levels similar to those at which strong inhibition was seen in paired-flash experiments. 5. Recovery from inhibition was not appreciably slowed when Ca2+ was injected to raise the free Ca2+ level between paired flashes. 6. We conclude that inhibition of InsP3-evoked Ca2+ liberation is not directly proportional to cytosolic free Ca2+ level and that recovery from inhibition in paired-pulse experiments involves factors other than the decline of cytosolic [Ca2+] following a conditioning response.

Animals↗
PubMed1994

Ca2+ influx modulation of temporal and spatial patterns of inositol trisphosphate-mediated Ca2+ liberation in Xenopus oocytes.

Inositol 1,4,5-trisphosphate (InsP3) functions as a second messenger by liberating intracellular Ca2+ and by promoting influx of extracellular Ca2+. We examined the effects of Ca2+ influx on the temporal and spatial patterns of intracellular Ca2+ liberation in Xenopus oocytes by fluorescence imaging of cytosolic free Ca2+ together with voltage clamp recording of Ca(2+)-activated Cl- currents. Oocytes were injected with a poorly metabolized InsP3 analogue (3-F-InsP3; see Introduction) to induce sustained activation of InsP3 signalling, and Ca2+ influx was controlled by applying voltage steps to change the driving force for Ca2+ entry. Positive- and negative-going potential steps (corresponding, respectively, to decreases and increases in Ca2+ influx) evoked damped oscillatory Cl- currents, accompanied by cyclical changes in cytosolic free Ca2+. The source of this Ca2+ was intracellular, since oscillations persisted when Ca2+ entry was suppressed by removing extracellular Ca2+ or by polarization close to the Ca2+ equilibrium potential. Fluorescence recordings from localized (ca 5 microns) spots on the oocyte showed repetitive Ca2+ spikes. Their frequency increased at more negative potentials, but they became smaller and superimposed on a sustained 'pedestal' of Ca2+. Spike periods ranged from about 50 s at +20 mV to 4s at potentials between -60 and -120 mV. Ca2+ spike frequency decreased after removing extracellular Ca2+, but the spike amplitude was not reduced and low frequency spikes continued for at least 30 min in the absence of extracellular Ca2+. Membrane current oscillations decayed in amplitude following voltage steps, while locally recorded Ca2+ spikes did not. This probably arose because Ca2+ release was initially synchronous across the cell, leading to large Ca(2+)-activated Cl- currents, but the currents then diminished as different areas of the cell began to release Ca2+ asynchronously. Fluorescence imaging revealed that Ca2+ liberation in 3-F-InsP3-loaded oocytes occurred as transient localized puffs and as propagating waves. Polarization to more negative potentials increased the frequency of puffs and the number of sites at which they were seen, and enhanced their ability to initiate waves. The frequency and velocity of Ca2+ waves increased at more negative potentials. When the potential was returned to more positive levels, repetitive Ca2+ spikes at first occurred synchronously across the recording area, but this synchronization was gradually lost and Ca2+ waves began at several foci. We conclude that influx of extracellular Ca2+ regulates the temporal and spatial patterns of Ca2+ liberation from InsP3-sensitive intracellular stores, probably as a result of dual excitatory and inhibitory actions of cytosolic Ca2+ on the InsP3 receptor.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

Metabolism of [3H]-noradrenaline in human dental pulp in vitro.

Slices of pulp from human maxillary and mandibular molar and promolar teeth were incubated with [3H]-noradrenaline (0.2 mumol/l) for 30 min after which the [3H]-noradrenaline and [3H]-metabolites in the tissue and medium were assayed by column chromatography. The deaminated metabolites 3,4-dihydroxy phenyl glycol (DOPEG) and 3,4-dihydroxy mandelic acid (DOMA) constituted 81% of the metabolites formed. Cocaine, an inhibitor of uptake1, decreased the formation of DOPEG and DOMA as well as the accumulation of [3H]-noradrenaline. In contrast to findings in rabbit pulp, when the disposition of exogenous noradrenaline in human pulp was examined by monoamine fluorescence histochemistry there was no evidence of extraneuronal accumulation of noradrenaline by connective tissue cells. In further experiments, pulp that had been incubated in [3H]-noradrenaline (0.6 mumol/l) for 30 min and superfused for 200 min contained [3H]-noradrenaline (183 pmol/g) and [3H]-DOMA (89 pmol/g). The 3H that overflowed into the perfusate between 85 and 90 min consisted mainly of metabolites. Stimulation of the sympathetic nerves through field electrodes increased the overflow of [3H]-noradrenaline into the perfusate threefold without affecting the overflow of metabolites. The increase was much greater (eightfold) in the presence of an alpha-adrenoceptor antagonist (rauwolscine; 0.1 mumol/l), plus inhibitors of uptake1 (desipramine; 0.3 mumol/l) and uptake 2 (corticosterone; 10 mumol/l). The results are interpreted as evidence that in human dental pulp the disposition of exogenous noradrenaline is determined largely by uptake by sympathetic nerves. After uptake, noradrenaline is deaminated by intraneuronal monoamine oxidase to DOPEG and DOMA.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Relation between intracellular Ca2+ signals and Ca(2+)-activated Cl- current in Xenopus oocytes.

Activation of inositol 1,4,5-trisphosphate (InsP3) signalling in Xenopus oocytes causes intracellular Ca2+ mobilization and thereby activates a Ca(2+)-dependent Cl- membrane conductance. Measurements of cytosolic Ca2+ levels using fluorescent indicators, however, revealed little correspondence with Cl- currents. Intracellular photorelease of InsP3 from a caged precursor evoked transient currents that peaked while the Ca(2+)-fluorescence signal was rising, and subsequently declined within a few seconds, even though the Ca2+ signal remained elevated much longer. Also, Cl- currents evoked by agonist activation showed transient spikes while a wave of Ca2+ liberation swept across the cell, but then decreased when the Ca2+ signal attained a maximal level. Thus, the Cl- current corresponded better to the rate of rise of intracellular free Ca2+, rather than to its steady state level. Experiments using paired flashes to photolyse caged InsP3 and caged Ca2+ indicated that this relationship did not arise through desensitization or inactivation of the Cl- conductance. Furthermore, fluorescence measurements made at different depths into the cell using a confocal microscope revealed no evidence that a rapid decline of local Ca2+ levels near the plasma membrane was responsible for the decay of Ca(2+)-activated Cl- current. Instead, Cl- channels may show an adaptive or incremental response to Ca2+, which is likely to be important for the encoding and transmission of information by Ca2+ spikes.

Animals↗