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R Nuccitelli

Publications and source records attributed to R Nuccitelli.

At least 37 records · Page 2Linked to original sources
PubMed1996

Fertilization stimulates an increase in inositol trisphosphate and inositol lipid levels in Xenopus eggs.

Previous experiments from our lab have suggested that the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) is required for sperm-induced egg activation in Xenopus laevis. Here we measure the endogenous production of both Ins(1,4,5)P3 and PIP2 during the sperm-induced and ionomycin-induced calcium wave in the egg and find that both increase following fertilization. Ins(1,4,5)P3 increases 3.2-fold from an unfertilized egg level of 0.13 pmole per egg (0.29 microM) to a peak of 0.42 pmole per egg (0.93 microM) as the calcium wave reaches the antipode in the fertilized egg. This continuous production of Ins(1,4,5)P3 during the time that the Ca2+ wave is propagating across the egg suggests the involvement of Ins(1,4,5)P3 in wave propagation. This increase in Ins(1,4,5)P3 is smaller in ionomycin-activated eggs than in sperm-activated eggs, suggesting that the sperm-induced production of Ins(1,4,5)P3 involves a PIP2 hydrolysis pathway that is not simply raising intracellular Ca2+. While one might expect PIP2 levels to fall as a result of hydrolysis, we find that PIP2 actually increases 2-fold. The total lipid fraction in unfertilized egg exhibits 0.8 pmole PIP2 per egg and this increases to 1.5 pmole as the calcium wave reaches the antipode. The PIP2 concentration peaks 2 min after the completion of the calcium wave at 1.8 pmole per egg. The amount of PIP2 in the animal and vegetal hemispheres of the egg was also measured by cutting frozen eggs in half. The vegetal hemisphere contained twice the amount of PIP2 as the animal hemisphere but it also contained twice the amount of lipid. Thus, there was an equivalent amount of PIP2 normalized to lipid in each hemisphere. Isolated animal and vegetal hemisphere cortices exhibit similar PIP2 concentrations, suggesting that the 2-fold higher total PIP2 in the vegetal half is not due to a gradient of PIP2 in the plasma membrane, but rather implies that cytoplasmic organelle membranes also contain PIP2.

Animals↗
PubMed1996

Imposition of a physiologic DC electric field alters the migratory response of human keratinocytes on extracellular matrix molecules.

Outwardly directed ionic currents have been measured leaving skin wounds in vivo. These currents generate physiologic electric fields of approximately 100 mV/mm, which may function to direct keratinocyte migration toward the healing wound. We investigated whether the substrate on which the keratinocyte migrates modulates the galvanotactic response to an electric migratory signal. Cultured human keratinocytes were plated on different matrices; types I and IV collagen, fibronectin, laminin, and tissue culture plastic. The effect of an applied direct current (DC) electric field on directional migration was monitored by time-lapse video microscopy over a 2-h period. Directionality was quantitated by calculating the cosine of the angle of migration in relation to anodal-cathodal orientation. Migration toward the negative pole was observed on all matrices as compared with controls (no applied field), which displayed random migration. No significant increase in directional response occurred when the field strength was increased by 100 mV/mm (physiologic levels) to 400 mV/mm. The degree of directionality and the average net cell translocation however, varied significantly with the substrate. The greatest cathodal migration in response to a DC electric field was observed with keratinocytes plated on types I and IV collagens and plastic. The directional migratory response was least on a laminin substrate, whereas cells on fibronectin demonstrated a response that was intermediate between those of collagen and laminin. These results suggest that physiologic ionic currents in concert with underlying matrix may influence the rate of reepithelialization of skin wounds.

Cell Movement↗
PubMed1996

Human keratinocytes migrate to the negative pole in direct current electric fields comparable to those measured in mammalian wounds.

Previous measurements of the lateral electric fields near skin wounds in guinea pigs have detected DC fields between 100-200 mV/mm near the edge of the wound. We have studied the translocation response of motile primary human keratinocytes migrating on a collagen substrate while exposed to similar physiological DC electric fields. We find that keratinocytes migrate randomly on collagen in fields of 5 mV/mm or less, but in larger fields they migrate towards the negative pole of the field, exhibiting galvanotaxis. Since these cells have an average cell length of 50 microns, this implies that they are able to detect a voltage gradient as low as 0.5 mV along their length. This cath-odally-directed movement exhibits increased directedness with increasing field strengths between 10 and 100 mV/mm. We observe a maximally directed response at 100 mV/mm with half of the cells responding to the field within 14 minutes. The average speed of migration tended to be greater in fields above 50 mV/mm than in smaller fields. We conclude that human keratinocytes migrate towards the negative pole in DC electric fields that are of the same magnitude as measured in vivo near wounds in mammalian skin.

Animals↗
PubMed1994

Highly polarized EGF receptor tyrosine kinase activity initiates egg activation in Xenopus.

Progesterone-matured Xenopus oocytes are arrested at second metaphase but resume meiosis following fertilization. To explore the role of tyrosine kinase activity and phosphatidylinositol turnover in this activation process, we caused oocytes to express three types of human epidermal growth factor receptor (EGF-R), which differ in their ability to stimulate these biochemical processes. Following mRNA injection we found that receptor expression was highly polarized, with most receptors located on the animal hemisphere. Occupancy of the wild-type EGF-R in progesterone-matured oocytes resulted in full egg activation as indicated by an activation potential, increased intracellular-free Ca2+ ([Ca2+]i), fertilization envelope liftoff, and cortical contraction. Fura-2 imaging showed that the wave of EGF-mediated Ca2+ release started in the animal hemisphere and progressed completely around the cell. These responses required receptor tyrosine kinase activity. Matured oocytes expressing the c'973 EGF-R, which possesses kinase activity but only weakly stimulates phosphatidylinositol turnover, responded differently to EGF addition. Cortical contraction and fertilization envelope liftoff appeared normal, but there was no activation potential. Significantly, [Ca2+]i was only slightly elevated and was topologically restricted to the regions expressing receptors. Our results suggest that some aspects of egg activation can occur through a tyrosine kinase pathway. However, phosphatidylinositol hydrolysis appears necessary for both amplification and propagation of signals generated locally by activated EGF-R.

Animals↗
PubMed1993

Protein kinases are required for embryonic neural crest cell galvanotaxis.

Embryonic quail neural crest cells migrate towards the negative pole of an imposed dc electric field as small as 7 mV/mm (0.4 mV per average cell length). The involvement of protein kinases in the mechanism utilized by these cells to detect and respond to such imposed fields was tested through the use of several kinase inhibitors. Evidence for the involvement of protein kinase C (PKC) included: (1) inhibition of the directed motility by 1 microM sphingosine that was reversed by the addition of the phorbol ester, PMA; (2) stimulation of a faster response to the imposed field by PMA; and (3) inhibition of the directed translocation by 5 microM H-7. However, another PKC inhibitor, staurosporine, did not inhibit the directed translocation (1 nM-1 microM). We also found evidence for the involvement of either cAMP- or cGMP-dependent protein kinase. The galvanotactic response was partially inhibited by the addition of 10 microM H-9 and the response was enhanced in the presence of the phosphodiesterase inhibitor, IBMX. However, the adenylate cyclase stimulant, forskolin, had no significant influence on the directed motility, although it reduced the average cell velocity. While these experiments suggest that cAMP- or cGMP-dependent protein kinase or PKC may be involved in the galvanotaxis response, two other protein kinases appeared not to be required. The myosin light chain kinase inhibitor, ML-7, had no effect on the directed motility in an imposed field, so myosin light chain kinase may not be required for galvanotaxis. Similarly, 5 microM W-7 had no significant effect on the directed translocation, suggesting that calmodulin-dependent protein kinase is not involved. Interestingly, the continuous activity of a protein kinase is apparently not required for the directed translocation response. The addition of the PKC and cAMP-dependent protein kinase inhibitor, H-7, after the cells had been exposed to the field for 1 hour, had no effect on the subsequent directed translocation. Thus, for these inhibitors to block the directed translocation, they must be present at the same time as the initial field application. This implies that an integral step in the cellular response mechanism for galvanotaxis involves the stimulation of a protein kinase whose effect is long lasting.

Animals↗
PubMed1993

The sperm-induced Ca2+ wave following fertilization of the Xenopus egg requires the production of Ins(1, 4, 5)P3.

We used fluorescence ratio imaging of fura-2 in the egg of Xenopus laevis to study the initiation and propagation of the wave of increased free Ca2+ that is normally triggered at fertilization. Naturally matured, jellied eggs were injected with fura-2 and ratio-imaged with fluorescence excitation at 350 and 385 nm while sperm were added. The [Ca2+]i rise normally begins as a small spot near the surface of the egg, remains fairly localized for 20-60 sec, and then spreads more rapidly across the egg at 7.5 +/- 0.05 microns/sec to reach the antipode about 5 min after fertilization. The [Ca2+]i wave velocity is slowed by increasing the concentration of fura-2 in the cytoplasm to 100 microM, and 250 microM fura-2 blocks wave propagation. The peak [Ca2+]i in a sperm-activated wave is 2.2 +/- 0.1 microM and [Ca2+]i returns to preactivation levels within 21 +/- 0.7 min after fertilization. We further studied the mechanism by which sperm trigger the Ca2+ wave by injecting substances that interfere with Ins(1,4,5)P3-induced Ca2+ release or Ca(2+)-induced Ca2+ release (CICR). Heparin (3 kDa) inhibits sperm-induced egg activation in a manner that is linearly proportional to its cytoplasmic concentration. At 130 microM (390 micrograms/ml), sperm-induced activation is completely blocked and at 75 microM (225 micrograms/ml) activation of half of the eggs is inhibited. All eggs injected with 130 microM heparin are polyspermic as verified using Hoechst dye to label nuclear DNA. Imaging eggs injected with 75 microM heparin revealed multiple, transient "spots" of increased [Ca2+]i that failed to spread across the egg. Injection of a monoclonal antibody to PIP2 (0.2 microM), kt3g, blocked sperm-induced egg activation in 73% of the 30 eggs injected, suggesting that activation requires the hydrolysis of PIP2 in the membranes of the egg rather than simply the introduction of Ins(1,4,5)P3 from the sperm. This sperm-induced egg activation is not blocked by either of two CICR inhibitors, procaine (10 mM) or ruthenium red (30 microM), and egg activation is not triggered by either of two stimulators of CICR, caffeine (10 mM) or ryanodine (50 microM).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Calcium buffer injections delay cleavage in Xenopus laevis blastomeres.

Microinjection of calcium buffers into the two-cell Xenopus laevis embryo delays cell division in a dose-dependent manner. Four calcium buffers in the BAPTA series with different affinities for calcium were used to distinguish between a localized calcium gradient regulating cleavage and the global calcium concentration regulating this event. DibromoBAPTA (Kd = 1.5 microM) was found to delay cleavage at the lowest intracellular concentration (1.3 mM) of the four buffers tested. The effectiveness of the calcium buffers was dependent upon the buffer dissociation constant but not in a linear fashion. The concentration of buffer required to delay cleavage increased as the buffer's dissociation constant shifted above or below that of the optimum buffer, dibromoBAPTA. This relationship between a calcium buffer's effectiveness at delaying cleavage and its calcium affinity provides support for the hypothesis that a calcium concentration gradient is required for normal cell cycle progression (Speksnijder, J. E., A. L. Miller, M. H. Weisenseel, T.-H. Chen, and L. F. Jaffe. 1989. Proc. Natl. Acad. Sci. USA. 86:6607-6611). DibromoBAPTA was also injected with two different amounts of coinjected calcium to test the possibility that the free calcium concentration of the buffer solution is the important parameter for delaying cleavage. However, we found that changes in buffer concentration have a much stronger effect than changes in the free calcium concentration. This observation supports the hypothesis that BAPTA-type buffers exert their effect by shuttling calcium from regions of high concentration to those of lower concentration, reducing any calcium concentration gradients present in the Xenopus embryo.

Animals↗
PubMed1993

Adrenergically activated Ca2+ increases in brown fat cells: effects of Ca2+, K+, and K channel block.

We measured intracellular calcium concentration ([Ca2+]i) during adrenergic stimulation using fura-2 ratio imaging of individual cultured neonatal rat brown fat cells. One micromolar norepinephrine (NE) increased [Ca2+]i from an average resting value of 105 nM to 555 nM in approximately 30 s. [Ca2+]i remained elevated as long as NE was present but returned to resting levels within 2-3 min after NE removal. The response was half maximal at approximately 50 nM NE and was primarily alpha-adrenergic. The sustained, but not the initial, increase in [Ca2+]i required extracellular calcium. Cells stimulated in high-K media had [Ca2+]i responses like those in 0 Ca2+, suggesting that depolarization abrogates calcium influx. Parallel perforated-patch recordings showed that the increase in [Ca2+]i activates a calcium-activated K conductance. Blocking K channels with moderate concentrations of tetraethylammonium (TEA) had only small effects on NE-induced changes in [Ca2+]i, but high concentrations of TEA significantly reduced the response. We conclude that cytoplasmic calcium is modulated by fluxes from both intracellular and extracellular sources and that K channels may not be required for normal short-term [Ca2+]i responses to hormone.

Adipose Tissue, Brown↗
PubMed1992

Inhibition of tyrosine phosphorylation prevents thrombin-induced mitogenesis, but not intracellular free calcium release, in vascular smooth muscle cells.

alpha-Thrombin, a G-protein-coupled receptor agonist, is mitogenic for neonatal vascular smooth muscle (VSM) cells, but it also causes secretion of the tyrosine kinase-coupled receptor agonist platelet-derived growth factor (PDGF). In order to determine the role of growth factors with tyrosine kinase-coupled receptors in thrombin's mitogenic signal transduction cascade, the synergistic effect of basic fibroblast growth factor (bFGF) in this system was examined. While bFGF itself is a growth factor for VSM cells, it causes a 1.7-fold synergistic effect when added together with thrombin. Herbimycin A, a specific tyrosine kinase inhibitor, both decreases thrombin-induced mitogenesis by greater than 90% and abolishes tyrosine phosphorylation of phospholipase C (PLC)-gamma-1. The magnitude and time course of the increase in intracellular free calcium concentration in response to thrombin is comparable in both the presence and absence of herbimycin A. These results provide evidence that herbimycin A specifically inhibits PLC-gamma-1 tyrosine phosphorylation without affecting VSM cell viability or calcium release. Furthermore, tyrosine phosphorylation is a necessary step in thrombin's mitogenic signal transduction cascade, but it is not essential for thrombin-induced release of calcium from intracellular stores. These data suggest that a tyrosine kinase, possibly supplied by the bFGF receptor, plays an essential role in thrombin-induced mitogenesis.

Animals↗
PubMed1992

Endogenous ionic currents and DC electric fields in multicellular animal tissues.

Through the use of the non-invasive vibrating probe technique for detecting extracellular ionic currents developed in 1974 [Jaffe and Nuccitelli: J Cell Biol 63:614-628, 1974], embryonic currents have been detected in a wide range of animal systems (recently reviewed in [Nuccitelli, Noninvasive Techniques in Cell Biology. New York: Wiley-Liss, 1990, pp 273-310]. In four of these studies, the corresponding electric field has been measured within the animal tissue. Such measurements of internal electric fields are quite challenging because they involve the insertion of microelectrodes into the developing tissue along specific regions of current flow. This paper reviews the evidence for endogenous transembryonic currents and dc electric fields in animal systems and provides the range of values for such physiological fields. These data should provide a guide to the range of imposed electric field strengths that could influence normal biological functions in living organisms.

Animals↗
PubMed1992

Inositol lipid hydrolysis contributes to the Ca2+ wave in the activating egg of Xenopus laevis.

We have used fluorescence ratio-imaging of fura-2 in the activating egg of Xenopus laevis to study the wave of increased intracellular free Ca2+ concentration ([Ca2+]i) while monitoring that of cortical granule exocytosis. Naturally matured eggs were dejellied, injected with fura-2, and activated by the iontophoresis of 1-30 nCoul of inositol-1,4,5-trisphosphate which triggers an immediate increase in free [Ca2+]i at the injection site. The Ca2+ rise spreads throughout the egg, reaching the opposite side in 5-8 min, and is followed by elevation of the fertilization envelope about 20-30 sec behind the [Ca2+]i wave. [Ca2+]i returns to preactivation levels within about 20 min after activation. We further studied the role of phosphatidylinositol-4,5-bisphosphate (PIP2) hydrolysis by microinjecting antibodies to PIP2 into the egg. PIP2 antibodies did not alter the propagation velocity of the wave but greatly reduced the amount of Ca2+ released in the egg cortex. These data suggest that PIP2 hydrolysis plays a role in the release of [Ca2+]i in the outer regions of the egg following activation.

Animals↗
PubMed1992

A computerized 2-dimensional vibrating probe for mapping extracellular current patterns.

We describe a computer-assisted 2-dimensional vibrating probe system for mapping endogenous electric current patterns in biological preparations. This system overcomes some of the main limitations of the original 1-dimensional vibrating probe design and adds several new capabilities. Two piezo-electric bender elements mounted perpendicularly are used to vibrate the probe in a circle by applying 2 sine waves (1 to each element) that are 90 degrees out-of-phase with each other. The circular rotation of the probe allows it to detect simultaneously the 2 orthogonal components of a current in the horizontal plane. The voltages measured by the probe are digitized and analyzed by a computer and are used to calculate a current vector. A graphical representation of the current vector is then superimposed on a video image of the experimental preparation. This probe system responds to known currents in the expected manner and exhibits a low inherent noise level. Also included in this paper are some preliminary measurements made with this instrument on neurulating Xenopus embryos and on transected larval sea lamprey (Petromyzon marinus) spinal cords.

Animals↗
PubMed1992

Development of functional calcium channels in cultured avian photoreceptors.

Vertebrate photoreceptors are unusual neurons in that they are capable of continuous calcium-mediated release of neurotransmitter (Trifonov, 1968; Hagins et al., 1970). In this study, we have examined the development and characteristics of calcium currents in chick cone cells placed in culture on embryonic day 8. Cone cells were identified by their lectin-binding properties, rhodopsin-like immunoreactivity, and the presence of an oil droplet. Using the whole-cell patch-clamp method, we have seen calcium currents in these cells after three days in culture, slightly before the appearance of synapses (Gleason & Wilson, 1989). Because cone calcium currents are blocked by cadmium and nifedipine but are enhanced by Bay K 8644, they most closely resemble L-type current (Nowycky et al., 1985). An unexpected feature of these currents is that their gating ranges varied widely between cells so that some cells showed the foot of their activation range at -70 mV and others as positive as -25 mV. Calcium imaging of fura-2 loaded cells was used to confirm the time course of calcium current development and describe the distribution of cytosolic calcium. As expected, depolarization of young cells failed to increase cytosolic calcium but in older cells an increase of threefold to fourfold was usually observed. Both at rest and during depolarization, most cone cells showed regional differences in internal calcium concentration. In the most mature cones, depolarization strongly elevated cytosolic calcium at the terminal end of the cell while producing a lesser change around the oil droplet and the ellipsoid region, suggesting that calcium channels are localized to the terminal.

Animals↗
PubMed1992

Reducing inositol lipid hydrolysis, Ins(1,4,5)P3 receptor availability, or Ca2+ gradients lengthens the duration of the cell cycle in Xenopus laevis blastomeres.

We have microinjected a mAb specifically directed to phosphatidylinositol 4,5-bisphosphate (PIP2) into one blastomere of two-cell stage Xenopus laevis embryos. This antibody binds to endogenous PIP2 and reduces its rate of hydrolysis by phospholipase C. Antibody-injected blastomeres undergo partial or complete arrest of the cell cycle whereas the uninjected sister blastomeres divided normally. Since PIP2 hydrolysis normally produces diacylglycerol (DG) and inositol 1,4,5-triphosphate (Ins[1,4,5]P3), we attempted to measure changes in the levels of DG following stimulation of PIP2 hydrolysis in antibody-injected oocytes. The total amount of DG in antibody-injected oocytes was significantly reduced compared to that of water-injected ones following stimulation by either acetylcholine or progesterone indicating that the antibody does indeed suppress PIP2 hydrolysis. We also found that the PIP2 antibodies greatly reduced the amount of intracellular Ca2+ released in the egg cortex during egg activation. As an indirect test for Ins(1,4,5)P3 involvement in the cell cycle we injected heparin which competes with Ins(1,4,5)P3 for binding to its receptor, and thus inhibits Ins(1,4,5)P3-induced Ca2+ release. Microinjection of heparin into one blastomere of the two-cell stage embryo caused partial or complete arrest of the cell cycle depending upon the concentration of heparin injected. We further investigated the effect of reducing any [Ca2+]i gradients by microinjecting dibromo-BAPTA into the blastomere. Dibromo-BAPTA injection completely blocked mitotic cell division when a final concentration of 1.5 mM was used. These results suggest that PIP2 turnover as well as second messenger activity influence cell cycle duration during embryonic cell division in frogs.

Animals↗
PubMed1991

Neural crest cell galvanotaxis: new data and a novel approach to the analysis of both galvanotaxis and chemotaxis.

The galvanotaxis response of neural crest cells that had migrated out of the neural tube of a 56-hr-old quail embryo onto glass coverslips was observed using time-lapse video microscopy. These cells exhibit a track velocity of about 7 microns/min and actively translocate toward the negative pole of an imposed DC electric field. This nonrandom migration could be detected for fields as low as 7 mV/mm (0.4 mV/cell length). We find that this directional migration is independent of the speed of migration and have generated a rather simple mathematical equation that fits these data. We find that the number of cells that translocate at a given angle, phi, with respect to the field is given by the equation N(phi) = exp(a0 + a1cos phi), where a1 is linearly proportional to the electric field strength for fields less than 390 mV/mm with a constant of proportionality equal to KG, the galvanotaxis constant. We show that KG = (150 mV/mm)-1, and at this field strength the cellular response is approximately half maximal. This approach to cellular translocation data analysis is generalizable to other directed movements such as chemotaxis and allows the direct comparison of different types of directed movements This analysis requires that the response of every cell, rather than averages of cellular responses, is reported. Once an equation for N(phi) is derived, several characteristics of the cellular response can be determined. Specifically, we describe 1) the critical field strength (390 mV/mm) below which the cellular response exhibits a simple, linear dependence on field strength (for larger field strengths, an inhibitory constant can be used to fit the data, suggesting that larger field strengths influence a second cellular target that inhibits the first); and 2) the amount of information the cell must obtain in order to generate the observed asymmetry in the translocation distribution (for a field strength of 100 mV/mm, 0.3 bits of information is required).

Animals↗
PubMed1991

The effects of inositol trisphosphates and inositol tetrakisphosphate on Ca2+ release and Cl- current pattern in the Xenopus laevis oocyte.

We report that Ins(1,3,4,5)P4 releases calcium from intracellular stores of intact Xenopus laevis oocytes, as indicated by two different techniques, Ca2(+)-sensitive microelectrodes and a fura-2 imaging system. Ins(1,3,4,5)P4 releases only 20% as much Ca2+ as the same amount of Ins(1,4,5)P3. This effect is not due to the conversion of the injected Ins(1,3,4,5)P4 to Ins(1,4,5)P3, which is known to release Ca2+, because the amount of [3H]Ins(1,3,4,5)P4 that is converted to Ins(1,4,5)P3 is extremely small, as determined using HPLC. Examination of the different current patterns induced by Ins(1,4,5)P3 and Ins(1,3,4,5)P4, when injected into voltage-clamped oocytes, provided further evidence that the Ins(1,3,4,5)P4 was not being converted back to Ins(1,4,5)P3. We investigated the effects of four compounds, three inositol trisphosphates (Ins(1,4,5)P3, Ins(2,4,5)P3, and Ins(1,3,4)P3), and Ins(1,3,4,5)P4, on Cl- current conductance in order to examine (1) the possible role of Ins(1,3,4,5)P4 in cell activation and (2) the relationships between intracellular Ca2+ and the activation of Cl- currents. Immature stage VI Xenopus laevis oocytes were voltage-clamped and injected with Ins(1,4,5)P3, Ins(2,4,5)P3, and Ins(1,3,4)P3. Ins(1,4,5)P3 and Ins(2,4,5)P3 triggered Ca2(+)-dependent Cl- currents, but Ins(1,3,4)P3 did not trigger currents nor did it release intracellular Ca2+. Ins(2,4,5)P3 was fourfold less effective at inducing the immediate Cl- current pulse than Ins(1,4,5)P3. The Cl- current pattern was quite dependent on the amount of Ins(1,4,5)P3 injected into the oocyte. Low amounts of Ins(1,4,5)P3 triggered only an immediate single Cl- current pulse, whereas large amounts triggered the immediate single pulse, followed by a quiescent period, followed by oscillating Cl- currents. In contrast to the response of Ins(1,4,5)P3, injection of Ins(1,3,4,5)P4 triggered only oscillating Cl- currents whose magnitude, but not pattern, was dependent on the amount injected into the cell. The currents generated by Ins(1,3,4,5)P4 resemble the oscillating Cl- currents triggered by large amounts of Ins(1,4,5)P3 and Ins(2,4,5)P3. Ins(1,3,4,5)P4, unlike Ins(1,4,5)P3 and Ins(2,4,5)P3, rarely caused an immediate Cl- current pulse, but caused an immediate release of calcium. Therefore, we suggest that the oscillating currents are only indirectly dependent on calcium. These [Ca2+]i and conductance measurements suggest that both Ins(1,4,5)P3 and Ins(1,3,4,5)P4 have roles in intracellular Ca2+ regulation.

Animals↗