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PubMed1998

Thimerosal-induced Ca2+ mobilization in isolated guinea pig cochlear outer hair cells.

Intracellular calcium mobilization of isolated guinea pig cochlear outer hair cells (OHCs) was investigated using thimerosal, a -SH group oxidizing agent, and fura-2 fluorescence ratio imaging microscopy. In the presence of thimerosal, intracellular Ca2+ concentrations ([Ca2+]i) of OHCs were elevated in a dose-dependent manner. Even in Ca(2+)-free medium, Ca2+ response was still induced. The effects of thimerosal on [Ca2+]i were completely blocked and reversed by dithiothreiotol (DTT). Neither 1-100 microM ryanodine nor 5-20 mM caffeine altered the effects of thimerosal. Pretreatment with pertussis toxin (PTX) for 30 min did not affect the thimerosal-induced increase in [Ca2+]i. The increase in [Ca2+]i when Ca2+ was added during thimerosal application in Ca(2+)-free medium was almost completely blocked by 500 microM LaCl3, while nifedipine did not inhibit further increase in [Ca2+]i caused by thimerosal. Thus, oxidation of the -SH group of the OHC membrane can induce a Ca2+ release from intracellular Ca2+ stores, which are ryanodine- and caffeine-insensitive, and Ca2+ influx through non-specific Ca2+ channels, but not the nifedipine-sensitive Ca2+ channels. The possible oxidation of -SH group gated Ca2+ channels in OHCs is worthy of further study.

Animals↗
PubMed2006

Thimerosal induces apoptosis in a neuroblastoma model via the cJun N-terminal kinase pathway.

The cJun N-terminal kinase (JNK)-signaling pathway is activated in response to a variety of stimuli, including environmental insults, and has been implicated in neuronal apoptosis. In this study, we investigated the role that the JNK pathway plays in neurotoxicity caused by thimerosal, an ethylmercury-containing preservative. SK-N-SH cells treated with thimerosal (0-10 microM) showed an increase in the phosphorylated (active) form of JNK and cJun with 5 and 10 microM thimerosal treatment at 2 and 4 h. To examine activator protein-1 (AP-1) transcription, cells were transfected with a pGL2 vector containing four AP-1 consensus sequences and then treated with thimerosal (0-2.5 microM) for 24 h. Luciferase studies showed an increase in AP-1 transcriptional activity upon thimerosal administration. To determine the components of the AP-1 complex, cells were transfected with a dominant negative to either cFos (A-Fos) or cJun (TAM67). Reporter analysis showed that TAM67, but not A-Fos, decreased AP-1 transcriptional activity, indicating a role for cJun in this pathway. To assess which components are essential to apoptosis, cells were treated with a cell-permeable JNK inhibitor II (SP600125) or transfected with TAM67, and the downstream effectors of apoptosis were analyzed. Cells pretreated with SP600125 showed decreases in activation of caspases 9 and 3, decreases in degradation of poly(ADP-ribose) polymerase (PARP), and decreased levels of proapoptotic Bim, in comparison to cells treated with thimerosal alone. However, cells transfected with TAM67 showed no changes in those same components. Taken together, these results indicate that thimerosal-induced neurotoxicity occurs through the JNK-signaling pathway, independent of cJun activation, leading ultimately to apoptotic cell death.

Anthracenes↗
PubMed1991

Vaccination despite thimerosal sensitivity.

Thimerosal sensitivity is extremely common in Austria, being surpassed as a contact allergen only by nickel. This high incidence is still rising and is probably due to the frequent vaccinations which are performed in Austria. Most of the patch-test-positive patients had recently been immunized with thimerosal-containing vaccines, and no other obvious sources of exposure to thimerosal could be found. On retrospective questioning, 48 out of 50 patients had had no problems with their recent immunization; the 2 who reported massive local reactions had received their injections, against the recommendation of the manufacturer, subcutaneously. In a prospective study, 12 thimerosal-sensitized persons received their follow-up immunization at our department, and no side effects occurred. This enables us to conclude that sensitization had occurred through vaccines, but that those amounts of thimerosal delivered i.m. are not sufficient to elicit clinical symptoms. Patch-test positivity to thimerosal thus represents no contra-indication to i.m. immunization with thimerosal-containing vaccines.

Adolescent↗
PubMed1994

Sensitization to thimerosal (Merthiolate) is still present today.

The results on thimerosal (Merthiolate) hypersensitivity of a retrospective study, together with the relevant data on thimerosal hypersensitivity referred to in the literature up to 1993, are presented. Positive patch test reactions to thimerosal (0.1% pet.) were observed in 32 (1.3%) of 2461 adult patients with suspected contact allergy examined in the period 1987-1992. 20 (0.8%) patients had a solitary positive patch test to thimerosal. The observed incidence is low. Clinical symptoms related to thimerosal hypersensitivity were observed in only 3 patients. The collected results are discussed with emphasis on the clinical implications of sensitization to thimerosal. It appears that a positive patch test to thimerosal is frequently clinically irrelevant.

Adult↗
PubMed1999

Sensitization to thimerosal in atopic children.

Thimerosal is an organic mercurial compound widely used as a preservative in vaccines, eyedrops, and contact lens cleaning and storage solutions. 5 infants, 2 female and 3 male, ranging in age from 7 to 28 months and affected by atopic dermatitis (AD) diagnosed according to the Hanifin and Rajka criteria, experienced an exacerbation of their clinical condition 2-10 days after mandatory vaccinations with vaccines containing thimerosal. Cutaneous lesions of nummular eczema appeared on the trunk, limbs and face. All patients were patch tested with serial dilutions of thimerosal in petrolatum. A positive patch test reaction to thimerosal 0.1% pet. was observed in all 5 children. 3 of them also showed a positive reaction at 0.01% and 0.05% pet. Despite their thimerosal-hypersensitivity, all children completed the entire series of mandatory vaccinations, care being taken to use different needles for injection and aspiration of the vaccine. The 2-year follow-up did not reveal other episodes of exacerbation of the AD after vaccination. The present study confirms the high frequency of sensitization to thimerosal in atopic children and suggests that vaccination can cause clinical symptoms in sensitized children. Nevertheless, sensitization to thimerosal does not prevent children from continuing with mandatory vaccinations.

Child, Preschool↗
PubMed1979

Human platelet aggregation by thimerosal. Functional and ultrastructural studies.

Thimerosal, a sulfhydryl group inhibitor, produces in an aggregometer a decrease in optical density of normal platelet-rich plasma over a wide range of concentrations. Ultrastructural study shows that the decrease of optical density produced by thimerosal at low doses is due to a true platelet aggregation preceded by a release reaction, whereas the aggregometric curves recorded after addition of thimerosal at high doses can be attributed to marked alterations of platelet morphology. Electron microscopic study shows the presence of electron-dense material between plasma membranes after addition of a low dose, and the early rupture of membranes after a high dose. These findings support previous conclusions that thimerosal binds to plasma membranes. Thimerosal induces a release reaction, seen in ultrastructural study and revealed by measurement of 14C-serotonin release. Moreover, thimerosal-induced aggregation is independent of released ADP and of formation of intermediates of the arachidonate pathway. Thimerosal-induced platelet aggregation is inhibited neither by ADP removal nor by aspirin addition.

Adenosine Diphosphate↗
PubMed1994

Thimerosal induces toxic reaction in non-sensitized animals.

The effects of injection of thimerosal solution on nonsensitized animals was investigated. Intrafootpad injection of thimerosal solution in nonsensitized mice resulted in a swelling response which peaked 1 h after injection and lasted for more than 24 h. Histopathological examination showed that there were severe edema and infiltration of polymorphonuclear neutrophils at the site of injection. An increased vascular permeability was observed after cutaneous injection of thimerosal solution on the back of nonsensitized rats. Since mercuric chloride and methyl mercury induced severer reactions, and thiosalicylic acid had no effect, mercury contained in thimerosal would have caused the reactions observed in this study. These results suggest that part of these hypersensitivity reactions against thimerosal observed among patients were possibly induced by the toxic effect of thimerosal. Therefore, thimerosal contained as a preservative in vaccine may augment the side-effects of the vaccination.

Animals↗
PubMed2005

In vitro uptake of glutamate in GLAST- and GLT-1-transfected mutant CHO-K1 cells is inhibited by the ethylmercury-containing preservative thimerosal.

Thimerosal, also known as thimersal, Merthrolate, or sodiumethyl-mercurithiosalicylate, is an organic mercurial compound that is used in a variety of commercial as well as biomedical applications. As a preservative, it is used in a number of vaccines and pharmaceutical products. Its active ingredient is ethylmercury. Both inorganic and organic mercurials are known to interfere with glutamate homeostasis. Brain glutamate is removed mainly by astrocytes from the extracellular fluid via high-affinity astroglial Na+-dependent excitatory amino acid transporters, glutamate/ aspartate transporter (GLAST) and glutamate transporter-1 (GLT-1). The effects of thimerosal on glutamate homeostasis have yet to be determined. As a first step in this process, we examined the effects of thimerosal on the transport of [3H]-d-aspartate, a nonmetabolizable glutamate analog, in Chinese hamster ovary (CHO) cells transfected with two glutamate transporter subtypes, GLAST (EAAT1) and GLT-1 (EAAT2). Additionally, studies were undertaken to determine the effects of thimerosal on mRNA and protein levels of these transporters. The results indicate that thimerosal treatment caused significant but selective changes in both glutamate transporter mRNA and protein expression in CHO cells. Thimerosal-mediated inhibition of glutamate transport in the CHO-K1 cell line DdB7 was more pronounced in the GLT-1-transfected cells compared with the GLAST- transfected cells. These studies suggest that thimerosal accumulation in the central nervous system might contribute to dysregulation of glutamate homeostasis.

Amino Acid Transport System X-AG↗
PubMed2003

Thimerosal and the occurrence of autism: negative ecological evidence from Danish population-based data.

OBJECTIVE: It has been suggested that thimerosal, a mercury-containing preservative in vaccines, is a risk factor for the development of autism. We examined whether discontinuing the use of thimerosal-containing vaccines in Denmark led to a decrease in the incidence of autism. DESIGN: Analysis of data from the Danish Psychiatric Central Research Register recording all psychiatric admissions since 1971, and all outpatient contacts in psychiatric departments in Denmark since 1995. PATIENTS: All children between 2 and 10 years old who were diagnosed with autism during the period from 1971-2000. OUTCOME MEASURES: Annual and age-specific incidence for first day of first recorded admission with a diagnosis of autism in children between 2 and 10 years old. RESULTS: A total of 956 children with a male-to-female ratio of 3.5:1 had been diagnosed with autism during the period from 1971-2000. There was no trend toward an increase in the incidence of autism during that period when thimerosal was used in Denmark, up through 1990. From 1991 until 2000 the incidence increased and continued to rise after the removal of thimerosal from vaccines, including increases among children born after the discontinuation of thimerosal. CONCLUSIONS: The discontinuation of thimerosal-containing vaccines in Denmark in 1992 was followed by an increase in the incidence of autism. Our ecological data do not support a correlation between thimerosal-containing vaccines and the incidence of autism.

Autistic Disorder↗
PubMed2000

[Thiol reagent thimerosal-induced Ca2+ mobilization in isolated guinea pig cochlear outer hair cells].

OBJECTIVE: To understand mechanism of cochlear outer hair cells (OHCs) intracellular Ca2+ mobilization further. METHODS: Intracellular calcium of isolated guinea pig was investigated using thimerosal, a--SH group oxidizing agent, and fura-2 fluorescence ratio imaging microscopy. RESULTS: In the presence of thimerosal, intracellular Ca2+ concentrations ([Ca2+]i) of OHCs were elevated in a dose-dependent manner. Even in Ca(2+)-free medium, Ca2+ response was still induced. The effects of thimerosal on [Ca2+]i were completely blocked and reversed by (DTT). Neither 1-100 mumol/L ryanodine nor 5-20 mmol/L caffeine altered the effects of thimerosal. Pretreatment with pertussis toxin (PTX) for 30 min did not affect the thimerosal-induced increase in [Ca2+]i The increase in [Ca2+]i when Ca2+ was added during thimerosal application in Ca(2+)-free medium was almost completely blocked by 500 mol/L LaCl3, while nifedipine did not inhibit further increase in [Ca2+]i caused by thimerosal. CONCLUSION: Oxidation of the -SH group of the OHC membrane can induce a Ca2+ release from intracellular Ca2+ stores, which are ryanodine- and caffeine-insensitive, and Ca2+ influx through non-specific Ca2+ channels, but not the nifedipine-sensitive Ca2+ Channels. The possible oxidation of--SH group gated Ca2+ channels in OHCs are worthy of further study.

Animals↗
PubMed1977

Effect of the ophthalmic preservative thimerosal on rabbit and human corneal endothelium.

Widespread use of the mercurial-containing preservative thimerosal as an antibacterial agent in ophthalmic drugs and solutions warranted an investigation into its possible cytotoxic effects on the functional and ultrastructural integrity of the corneal endothelium. No changes in corneal thickness were observed during 5 hours' perfusion of the endothelium of rabbit and human corneas with 0.0001 and 0.0005 percent thimerosal in glutathione bicarbonate Ringer's solution (GBR). Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) of the endothelium of the 0.0001 percent group revealed normal ultrastructure. SEM and TEM of the endothelium of corneas perfused with 0.0005 percent thimerosal for 5 hours revealed condensed mitochondria, cytoplasmic vacuoles, and cytoplasmic flaps at the apical end of the cellular junctions. Perfusion of higher concentrations (0.001 and 0.005 perecnt) of thimerosal in GBR resulted in increases in corneal thickness after 2 hours and irreversible ultrastructural damage to the endothelial cells by 5 hours. Corneas perfused with 0.01 and 0.1 percent thimerosal in GBR showed a rapid and immediate increase in corneal thickness and endothelial cell death and necrosis within 1 hour. It is postulated that the mercury in thimerosal becomes bound to the cell membrane protein sulfhydryl groups, causing an increase in cellular permeability; These results suggest that the prolonged exposure of the corneal endothelium to thimerosal in the accepted antimicrobial dosage of 0.005 to 0.001 percent may result in functional and structural damage to the endothelium.

Animals↗
PubMed1981

Specific modification of gastric K+-stimulated ATPase activity by thimerosal.

Treatment of hog gastric microsomes with the sulfhydryl reagent, thimerosal (ethylmercurithiosalicylate), produced differential effects on the K+-ATPase and the K+-stimulated p-nitrophenylphosphatase activities. For example, exposure to 2 mM thimerosal for 3 min severely reduced the activity of K+-stimulated ATPase, while K+-p-nitrophenylphosphatase activity was enhanced 2- to 3-fold. Higher concentration of thimerosal, or longer incubation times, also led to inhibition of K+-p-nitrophenylphosphatase. The activated state of p-nitrophenylphosphatase could be sustained by a 20-fold, or greater, dilution of treated membranes, and could be reversed by reduction of membrane SH groups by exogenous thiols. Significant activation of K+-p-nitrophenylphosphatase was not produced by p-chloromercuribenzene sulfonate, p-chloromercuribenzoate or mersalyl; however, ethyl mercuric chloride had qualitatively similar activity effects as thimerosal. Kinetics of K+-p-nitrophenylphosphatase for thimerosal-treated membranes were altered as follows: V increased; Km for p-nitrophenylphosphate unchanged for Ka for K+ increased. ATP, which is a potent inhibitor of K+-p-nitrophenylphosphatase activity in native membranes (KI approximately 200 microM). These data suggest that there are multiple SH groups which differentially influence the gastric K+-stimulated ATPase activity. Defined treatments with thimerosal are interpreted as an uncoupling of the K+-stimulated phosphatase component of the enzyme (for which p-nitrophenylphosphatase is a presumed model reaction). Such differential modifications can be usefully applied to the study of partial reactions of the enzyme and their specific role in the related H+-transport reaction.

4-Nitrophenylphosphatase↗
PubMed1989

The sulfhydryl reagent thimerosal elicits human platelet aggregation by mobilization of intracellular calcium and secondary prostaglandin endoperoxide formation.

The effect of the sulfhydryl (SH) group inhibitor ethylmercurithiosalicylate (thimerosal) on the function of human platelets was investigated. In contrast to known SH reagents such as p-chloromercuribenzoate or N-ethylmaleimide, thimerosal elicited both aggregation and [3H]serotonin release of washed human platelets at low micromolar concentrations (greater than or equal to 2 microM). Only a significant higher dose (greater than or equal to 15 microM) was effective when platelets were pretreated with the cyclooxygenase inhibitor aspirin, indicating an amplification of the proaggregatory effect of thimerosal by secondary prostaglandin (PG) endoperoxide and/or thromboxane (TX) formation. Consistent with this notion, thimerosal induced endogenous platelet arachidonic acid (20:4) metabolism which could be attributed to enhanced 20:4 liberation, presumably by activation of phospholipase A2. The latter effect was mediated by mobilization of intracellular calcium (Ca2+), and was not affected by removal of extracellular Ca2+. In the presence of aspirin, the thimerosal-induced Ca2+ elevation was completely reversed by dithiothreitol (DTT) which implicates SH groups in intracellular Ca2+ transport. In contrast to previous observations with other SH reagents, thimerosal had no effect on the inositoltrisphosphate (IP3)-mediated release or the sequestration (and/or extrusion) of intracellular Ca2+ following stimulation with thrombin, indicating an action on an as yet undefined CA2+ transport system.

Aspirin↗
PubMed2004

Effect of thimerosal, a preservative in vaccines, on intracellular Ca2+ concentration of rat cerebellar neurons.

The effect of thimerosal, an organomercurial preservative in vaccines, on cerebellar neurons dissociated from 2-week-old rats was compared with those of methylmercury using a flow cytometer with appropriate fluorescent dyes. Thimerosal and methylmercury at concentrations ranging from 0.3 to 10 microM increased the intracellular concentration of Ca2+ ([Ca2+]i) in a concentration-dependent manner. The potency of 10 microM thimerosal to increase the [Ca2+]i was less than that of 10 microM methylmercury. Their effects on the [Ca2+]i were greatly attenuated, but not completely suppressed, under external Ca(2+)-free condition, suggesting a possibility that both agents increase membrane Ca2+ permeability and release Ca2+ from intracellular calcium stores. The effect of 10 microM thimerosal was not affected by simultaneous application of 30 microM L-cysteine whereas that of 10 microM methylmercury was significantly suppressed. The potency of thimerosal was similar to that of methylmercury in the presence of L-cysteine. Both agents at 1 microM or more similarly decreased the cellular content of glutathione in a concentration-dependent manner, suggesting an increase in oxidative stress. Results indicate that thimerosal exerts some cytotoxic actions on cerebellar granule neurons dissociated from 2-week-old rats and its potency is almost similar to that of methylmercury.

Animals↗
PubMed2001

Modulation of inositol 1,4,5-trisphosphate binding to the various inositol 1,4,5-trisphosphate receptor isoforms by thimerosal and cyclic ADP-ribose.

Three different genes encode the inositol 1,4,5-trisphosphate (IP3) receptor (IP3R), an intracellular Ca2+ channel involved in cellular Ca2+ signaling. The IP3-binding characteristics of the various IP3R isoforms differ, but until now no specific activators or inhibitors of IP3 binding have been described. We compared the effects of oxidizing reagents, in particular thimerosal, and of cyclic ADP-ribose (cADPR) on IP3 binding to the various IP3R isoforms. We therefore expressed the N-terminal 581 amino acids of the three IP(3)R isoforms as recombinant proteins in the soluble fraction of Escherichia coli (ligand-binding sites [lbs] 1, 2, and 3) as well as the full-length IP3R1 and IP3R3 in Spodoptera frugiperda (Sf9) insect cells. Thimerosal (100 microM) stimulated IP3 binding to lbs-1 (1.4-fold) and lbs-3 (2.5-fold), but had no effect on lbs-2. Thimerosal acted on lbs-1 and lbs-3 by decreasing the Kd for IP3 binding (from 46 +/- 4 nM to 20 +/- 2 nM and from 54 +/- 21 nM to 19 +/- 7 nM for lbs-1 and -3, respectively) without modifying the Bmax. Similarly, IP3 binding to microsomes of Sf9 insect cells overexpressing the full-length IP3R1 was 1.2-fold stimulated by thimerosal. Thimerosal, however, did not affect IP3 binding to Sf9-IP3R3 microsomes, suggesting that in situ thimerosal will only directly affect ligand binding to the type 1 isoform. cADPR (50 microM) stimulated IP3 binding to Sf9-IP3R1 microsomes (1.5-fold), but not to Sf9-IP3R3 microsomes. In addition, cADPR inhibited IP3 binding to lbs-1 and lbs-2 by decreasing the affinity for IP3 1.8- and 2.8-fold, respectively, while IP3 binding to lbs-3 was not affected. These results suggest that a regulatory site for cADPR is present in the ligand-binding domain of IP3R1 and 2, but not of IP3R3.

Adenosine Diphosphate Ribose↗
PubMed2002

Thimerosal stimulates focal adhesion kinase and cytoskeletal changes by redox modulation.

Thimerosal is one of the most widely used preservatives and has been reported to cause chemically mediated side effects. However, the mechanism of the side effects is not clearly understood yet. In the present study, we showed that HeLa S cells treated by thimerosal generated reactive oxygen species (ROS). Thimerosal-generated ROS stimulated the tyrosine phosphorylation of focal adhesion kinase (FAK) and also induced cytoskeletal changes. Pretreatment with intracellular calcium chelator, BAPTA did not block the thimerosal-mediated FAK tyrosine phosphorylation. On the other hand, either FAK inhibitor, tyrphostin or ROS scavenger, N-acetyl-L-cysteine (NAC) suppressed the tyrosine phosphorylation and cytoskeletal changes. These results suggest that thimerosal seems to induce FAK tyrosine phosphorylation and cytoskeletal changes by ROS generation but not by intracellular calcium mobilization. We think the present finding can be an important clue to understanding the mechanism of thimerosal-mediated side effects, such as contact dermatitis, and allergy.

Acetylcysteine↗
PubMed1999

Time course of cortical and zona reactions of pig oocytes upon intracellular calcium increase induced by thimerosal.

Our previous study indicated that thimerosal is one of the most effective artificial activators to mimic sperm-induced increases in the intracellular free calcium concentration ([Ca2+]i) and other activation events in pig oocytes (Macháty et al., 1997). The present study was conducted to examine the temporal relationship between intracellular calcium transients, cortical granule (CG) exocytosis and the zona reaction induced by thimerosal. When pig oocytes matured in vitro were exposed to 200 microM thimerosal the first intracellular calcium transient, with a mean peak ratio of 4.97 +/- 1.14, was observed 509.64 +/- 122.03 s after addition of thimerosal. The density of CGs fell significantly from 63.3 +/- 11.7 CGs/100 micron 2 of cortex in control oocytes to 25.7 +/- 19.2 CGs/100 micron 2 of cortex (59.4% release) at 2 min after the first intracellular calcium transient. At 5 min after the calcium transient the residual CG density had been reduced to 10.7 +/- 10.4 CGs/100 micron 2 of cortex (83.1% release). This degree of CG exocytosis was the same as that in oocytes penetrated by sperm (9.5 +/- 5.1 CGs/100 micron 2 of cortex). No further decrease in residual CG density was observed at 10 min (10.3 +/- 14.8 CGs/100 micron 2 of cortex). Whereas 77.4% (120/155) of control oocytes were penetrated by spermatozoa only 1.4% (2/144) of thimerosal-treated oocytes were penetrated. Further experimental results obtained by in vitro fertilisation of oocytes with preincubated (capacitated) spermatozoa suggested that the zona block to sperm penetration in thimerosal-treated oocytes occurred within 35 min after CG exocytosis and 40 min after the first calcium transient. These results indicate that polyspermic penetration of pig oocytes inseminated in vitro is not due to delayed or incomplete CG exocytosis but more likely to a delayed zona reaction and/or simultaneous sperm penetration.

Animals↗
PubMed1997

Effects of thimerosal on the transient kinetics of inositol 1,4,5-trisphosphate-induced Ca2+ release from cerebellar microsomes.

Thimerosal, a thiol-reactive reagent, has been shown to increase the cytosolic Ca2+ concentration in a variety of cells by sensitizing inositol 1,4,5-trisphosphate (InsP3) receptors. Thimerosal can have both sensitizing (at concentrations of <2 microM) and inhibitory (at concentrations of >2 microM) effects on InsP3-induced Ca2+ release (IICR) from cerebellar microsomes. Transient kinetic studies were performed by employing a fluorimetric stopped-flow approach using fluo-3. IICR was found to be a bi-exponential process with a fast and a slow component. At a maximal InsP3 concentration (20 microM), the fast phase had a rate constant of 0.9 s-1 and the slow phase had a rate constant of 0.4 s-1. The amplitudes of the two phases were 60% and 40% respectively. When the rate constants for the two phases were plotted as Hill plots, the processes were found to be non-co-operative in both cases (Hill coefficient of 1.0), thus arguing for a simple mechanism linking InsP3 binding to channel opening. At a submaximal InsP3 concentration (0.2 microM), where the sensitizing effects of thimerosal are most pronounced, thimerosal increased the rate constants of both phases in a sigmoidal fashion, with a Hill coefficient of 4.0, suggesting that several cysteine residues (up to four) need to be modified in order for maximum sensitization to occur. The rate constants remained elevated even at thimerosal concentrations that inhibited IICR. The amplitude or extent of Ca2+ release was, however, elevated to a much greater extent in the slow phase, suggesting that the two phases respond differently. At maximal InsP3 concentrations, thimerosal has no effect upon the rate constants but inhibits the amplitude of Ca2+ release.

Animals↗