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J B Lombardini

Publications and source records attributed to J B Lombardini.

At least 19 recordsLinked to original sources
PubMed1992

Effects of taurine on the phosphorylation of specific proteins in subcellular fractions of the rat retina.

The effects of 20 mM taurine on the phosphorylation of specific proteins in mitochondrial and rod outer segment subcellular fractions of the rat retina were measured. A band of protein with an apparent molecular weight of approximately 20K was consistently inhibited by taurine. Densitometry measurements performed on gel electrophoresis autoradiograms from the mitochondrial fraction demonstrated a 42.7 +/- 8.3% decrease due to taurine (20 mM) in the area corresponding to radioactivity from the approximately 20K phosphoprotein. However, only a 21.2 +/- 9.0% decrease was observed due to taurine in the rod outer segment preparation. These data suggest that taurine is exerting its primary effect on the phosphorylation of the approximately 20K molecular weight protein in the mitochondria of the retina. In addition, calmodulin and phorbol ester had no effect on the phosphorylation of the approximately 20K molecular weight protein.

Animals↗
PubMed1991

Inhibition by taurine of the phosphorylation of specific synaptosomal proteins in the rat cortex: effects of taurine on the stimulation of calcium uptake in mitochondria and inhibition of phosphoinositide turnover.

It has been previously observed that taurine inhibits PKC-activated phosphorylation of specific proteins including a approximately 20k Mr protein in rat cortical synaptosomes. In the present study, the mechanism of the above effects of taurine were investigated. In an intrasynaptosomal cytosol fraction obtained by subcellular fractionation, taurine did not have inhibitory effects on protein phosphorylation. However, taurine did inhibit the phosphorylation of the approximately 20k Mr protein in a reconstituted preparation containing intrasynaptosomal cytosol and mitochondria. Experiments measuring calcium uptake demonstrated that taurine increased the accumulation of 45Ca2+ in the mitochondrial fraction in incubation systems both in the absence and presence of added ATP. In addition, taurine inhibited the accumulation of 32P-labeled phosphatidic acid in synaptosomes indicative of a reduction in the levels of diacylglycerol. These results suggest that taurine may inhibit specific protein phosphorylation both by reducing cytosolic calcium levels and by inhibiting the turnover of phosphoinositides. These effects of taurine on the signal transduction cascade involving PKC and phosphoinositide metabolism indicate a potential biological role for taurine in the nervous system.

Adenosine Triphosphate↗
PubMed1991

Taurine inhibits protein kinase C-catalyzed phosphorylation of specific proteins in a rat cortical P2 fraction.

We previously reported that taurine inhibits the phosphorylation of specific proteins in a P2 synaptosomal fraction prepared from the rat cortex. In the present study, the regulation of the phosphorylation of an approximately 20K Mr protein whose phosphorylation is inhibited by taurine was further investigated. The phosphorylation of the approximately 20K Mr protein in a hypo-osmotically shocked P2 fraction from rat cortex was dependent on the free Ca2+ in the reaction medium. Depolarization induced by 30 mM K+ stimulated the phosphorylation of the approximately 20K Mr protein in an intact synaptosomal P2 preparation by 30-fold. This stimulation was inhibited 35% by taurine, whereas guanidinoethanesulfonic acid, a taurine analogue, did not have any effect, thereby indicating the specificity of taurine. Addition of phorbol 12-myristate 13-acetate, a phorbol ester, together with phosphatidylserine, stimulated the phosphorylation of the approximately 20K Mr protein in the hypo-osmotically shocked P2 synaptosomal fraction by fivefold, whereas cyclic AMP, cyclic GMP, and calmodulin did not have any effect on the phosphorylation of this particular protein. Phorbol 12-myristate 13-acetate-stimulated phosphorylation of the approximately 20K Mr protein is blocked 30% by taurine. Taurine also inhibited phorbol 12-myristate 13-acetate-activated phosphorylation of two other proteins that were similar in molecular weight and isoelectric point to the approximately 20K Mr protein on two-dimensional gels. These results suggest that taurine modulates the phosphorylation of specific proteins regulated by the signal transduction system in the brain. Thus, taurine may modulate neuroactivity by inhibiting the phosphorylation of specific proteins involved in regulatory function.

Animals↗
PubMed1990

Taurine analogues as modifiers of the accumulation of 45calcium ions in a rat retinal membrane preparation.

The effects of a series of taurine analogues on the accumulation of 45calcium ions in a crude rat retinal membrane preparation are reported. 45Calcium ion accumulation was measured at high (1.4 mM) calcium ion concentration in the presence and absence of ATP. In the absence of ATP, taurine and alpha-sulfo-beta-alanine both inhibit 45calcium ion accumulation. alpha-Sulfo-beta-alanine is the more potent of the two compounds (50% vs. 30% inhibition at 20 mM). The trans aminocycloalkanesulfonic acid analogues of taurine [(+/-) trans-2-aminocyclopentanesulfonic acid and (+/-) trans-2-aminocyclohexanesulfonic acid] are stimulators (3- to 4-fold) of 45calcium ion accumulation. In the presence of ATP (1.2 mM), taurine and the analogues of taurine had no effect. Structure-activity-relationships of these compounds pertinent to their effects on 45calcium ion accumulation at a high calcium ion concentration and in the absence of ATP are discussed. In addition, the inhibitory effects of taurine on 45calcium accumulation in subfractions of the retina (rod outer segments, synaptosomes, and mitochondria) and the effects of the divalent ionophore A23187 on 45calcium accumulation were also investigated.

Adenosine Triphosphate↗
PubMed1990

Inhibitory and stimulatory effects of structural and conformational analogues of taurine on ATP-dependent calcium ion uptake in the rat retina: deductions concerning the conformation of taurine.

A number of novel analogues of taurine were tested in a rat retinal preparation for their stimulatory or inhibitory activity in the ATP-dependent calcium ion uptake system at low calcium ion concentration. While the structural requirements for maintaining biological activity were quite limited, certain sulfone derivatives of taurine were observed to be more potent stimulators of calcium ion uptake than taurine. Utilization of multiple effectors in the uptake system was also analyzed. The effects of all combinations of taurine plus analogue that were tested were demonstrated to be mutually exclusive, that is, the compounds have similar modes of action. When two inhibitors, TAPS and THQS, were tested in combination, kinetic evaluation of the data suggested that both of these compounds also have a similar mode of action (mutually exclusive) with respect to each other. The inhibitors, TAPS and THQS, were also observed to be non-competitive with respect to taurine thus suggesting that they do not bind to the same site as taurine.

Adenosine Triphosphate↗
PubMed1989

Sulfone analogues of taurine as modifiers of calcium uptake and protein phosphorylation in rat retina.

The syntheses of five sulfone analogues of taurine are described: 2-aminoethylmethyl sulfone (AEMS), thiomorpholine-1,1-dioxide (TMS), N-methylthiomorpholine-1,1-dioxide (M-TMS), (+/-)3-aminotetrahydrothiopyran-1,1-dioxide (APS), and (+/-)3-aminotetrahydrothiophene-1,1-dioxide (ATS). When these compounds were evaluated in the rat retina as modulators of ATP-dependent calcium ion uptake at low calcium ion concentrations (10 microM), AEMS, ATS, and APS were found to be more potent stimulators of ATP-dependent calcium ion uptake than taurine. TMS and M-TMS had no effect. At high concentrations of calcium ions (1.44 mM), taurine, AEMS, ATS, APS, and TMS inhibited ATP-independent calcium ion uptake; AEMS, ATS, and APS were more potent inhibitors than taurine. ATS was the only compound tested (including taurine) that inhibited ATP-dependent calcium ion uptake at high calcium ion concentrations. The effects of the sulfone analogues of taurine on the incorporation of phosphate into retinal proteins were also studied. Taurine, AEMS, ATS, APS, and TMS were equipotent inhibitors of phosphate incorporation (30-45%). M-TMS had no effect.

Adenosine Triphosphate↗
PubMed1989

Analogues of taurine as stimulators and inhibitors of ATP-dependent calcium ion uptake in rat retina: combination kinetics.

Taurine is an amino acid that plays important roles in maintaining both the structural integrity and function of the retina. Thus, the effects of taurine, taurine analogues, and their combinations were studied in the ATP-dependent calcium ion uptake system at low calcium ion concentrations (10 microM) in a rat retinal membrane preparation. (+/-)-(trans)-2-Aminocyclopentanesulfonic acid (TAPS), a cyclic taurine analogue previously determined to inhibit ATP-dependent calcium ion uptake was demonstrated to be noncompetitive (Ki = 0.055 mM) with respect to taurine, that is, the values for the half-saturation concentrations calculated from varying concentrations of taurine compared with varying concentrations of taurine in the presence of a fixed concentration of TAPS (80 microM) did not change. However, the values for the maximal rates of change were significantly different. 1,2,3,4-Tetrahydroquinoline-8-sulfonic acid (THQS), a less potent inhibitor of ATP-dependent calcium ion uptake than TAPS, was also shown to be noncompetitive with taurine, with an inhibition constant (Ki) of 23.8 mM. Thus, it is presumed that both compounds (TAPS and THQS) are acting at receptor site(s) other than the taurine binding site. When TAPS and THQS were tested in a mixture that maintains a ratio (fixed ratio mixture) of 1 part TAPS and 25 parts THQS (by concentration, in mM), varied over a wide range of concentrations, and were then analyzed by median-effect plots and equation, the inhibitory effects are strongly synergistic, as shown by the combination index and the dose-reduction index. The parallel nature of the median-effect plots of TAPS and THQS indicates that the two inhibitors have a similar mode of action, that is, mutually exclusive. (+/-)-3-Aminotetrahydrothiophene-1,1-dioxide (ATS) and (+/-)-piperidine-3-sulfonic acid (PSA) are an agonist and partial agonist that demonstrated stimulatory effects on ATP-dependent calcium ion uptake. When tested in combination (1:1) with taurine, they were also determined to be mutually exclusive. It was demonstrated that ATS and taurine induced the same maximal rates of change of calcium ion uptake; however, PSA was less potent than taurine. The combination of taurine plus ATS was additive, whereas the combination of taurine plus PSA was synergistic. Structure-activity relationships of the taurine analogues and their topological relationships are discussed.

Adenosine Triphosphate↗
PubMed1988

Effects of aminocycloalkanesulfonic acid analogs of taurine on ATP-dependent calcium ion uptake and protein phosphorylation.

A cyclopentane analog of taurine [(+/-)cis-2-aminocyclopentanesulfonic acid] (CAPS) was synthesized, and its effects on ATP-dependent calcium ion uptake and protein phosphorylation in rat retina were investigated along with other cyclic analogs of taurine, (+/-)trans-2-Aminocyclopentanesulfonic acid (TAPS) is the most potent aminocycloalkanesulfonic acid inhibitor of ATP-dependent calcium ion uptake in retinal homogenates [S.M. Leibowitz, J.B. Lombardini and P.S. Salva, Biochem. Pharmac. 36, 2109 (1987)], eliciting its effects in the micromolar range (I50 = 39 +/- 5 microM). CAPS was found to be a less potent aminocycloalkanesulfonic acid inhibitor (I50 = 1780 +/- 400 microM) of ATP-dependent calcium ion uptake in retinal homogenates. Taurine inhibited phosphate incorporation into rat retinal proteins, whereas TAPS, TAHS and CAPS stimulated incorporation.

Adenosine Triphosphate↗
PubMed1988

Effects of taurine and mitochondrial metabolic inhibitors on ATP-dependent Ca2+ uptake in synaptosomal and mitochondrial subcellular fractions of rat retina.

ATP-dependent Ca2+ uptake was investigated at low Ca2+ concentrations (10 microM) in rat retinal synaptosomal and mitochondrial preparations obtained by differential centrifugation on Ficoll gradients. Ca2+ uptake in the synaptosomal and mitochondrial subcellular preparations was stimulated by ATP and additionally stimulated by ATP plus taurine. The ATP-dependent and taurine-stimulated ATP-dependent Ca2+ uptakes were inhibited by mitochondrial metabolic inhibitors (atractyloside, oligomycin, and ruthenium red). These metabolic inhibitors had a greater effect on the ATP-dependent and taurine-stimulated ATP-dependent Ca2+ uptake activities in the mitochondrial preparation than in the synaptosomal preparation. ATP-dependent Ca2+ uptake in a synaptosomal subfraction obtained by osmotic shock was only partially inhibited by atractyloside. ATP-dependent Ca2+ uptake in the synaptosomal subfraction was also stimulated by taurine but to a lesser extent than in either the synaptosomal or mitochondrial preparation. These studies suggest that mitochondria are primarily responsible for taurine-stimulated ATP-dependent Ca2+ uptake in synaptosomal preparations.

Adenosine Triphosphate↗
PubMed1987

Cyclic taurine analogs. Synthesis and effects on ATP-dependent Ca2+ uptake in rat retina.

Syntheses of (+/-)trans- and (+/-)cis-2-aminocyclohexane sulfonic acid (TAHS and CAHS) and (+/-)trans-2-aminocyclopentane sulfonic acid (TAPS) were achieved. In solution, the preferred conformations of TAHS and CAHS have been determined by high field NMR to be diequatorial and equatorial (sulfonic acid moiety)-axial (amino moiety) respectively. When these agents were evaluated as cyclic analogs of taurine in rat retina, TAHS and TAPS inhibited ATP-dependent calcium uptake in the micromolar range, whereas CAHS stimulated calcium uptake in the millimolar range in a manner similar to taurine. TAHS and TAPS are the most potent inhibitors of ATP-dependent calcium uptake in the rat retinal preparation yet reported.

Adenosine Triphosphate↗
PubMed1987

A model of the compartmentalization of taurine in rat hypothalmaic neuronal and glial cell particles.

We conclude that the high affinity taurine uptake system is located on neuronal membranes in the hypothalamus and that the low affinity taurine uptake system is located on glial membranes. The evoked release of taurine by either K+ and/or veratridine is Ca2+-independent and thus taurine, according to classical criteria, does not function as a neurotransmitter in the hypothalamus. This model for the differential cellular transport and compartmentalization of taurine into neuronal and glial cells and for a cytoplasmic location has important implications concerning its possible role in the central nervous system, specifically as a neuromodulator of neuronal activity.

Animals↗
PubMed1987

The relationship between sodium and high-affinity taurine uptake in hypothalamic crude P2 synaptosomal preparations.

Two uptake systems for taurine transport in a rat hypothalamic crude synaptosomal preparation were identified. The true transport constants were, for the high-affinity uptake system, Km = 240 microM and V (maximum velocity) = 400 nmol/g protein/min, and for the low-affinity uptake system, Km = 5290 microM and V = 1640 nmol/g protein/min. The initial velocity of high-affinity taurine uptake by the crude synaptosomal preparation was studied as a function of sodium and taurine concentration. Hill plots were constructed from these data. The requirement of high-affinity taurine uptake on a sodium gradient was examined by utilizing monensin, and the metabolic poisons, 2,4-dinitrophenol and ouabain. The major findings are as follows: 1) One sodium ion is co-transported with each taurine molecule; 2) the high-affinity uptake process is driven by the sodium concentration gradient across the membrane; 3) sodium increases the maximal velocity rather than the affinity of the high-affinity taurine carrier for the taurine molecule; 4) one taurine molecule is transported per carrier for both the high- and low-affinity taurine uptake systems; and 5) high-affinity taurine uptake is an energy-dependent process.

2,4-Dinitrophenol↗
PubMed1987

Is taurine a hypothalamic neurotransmitter?: A model of the differential uptake and compartmentalization of taurine by neuronal and glial cell particles from the rat hypothalamus.

Although taurine has been postulated to be a neurotransmitter or neuromodulator in the mammalian CNS, little is known concerning its role in brain function. Evidence suggesting that taurine may influence endocrine and homeostatic mechanisms via the hypothalamus resulted in our investigations into its function in this brain region. The main objectives of the research were to characterize the specific binding, uptake, and release of taurine in the hypothalamus. A specific aim was to examine the proposed neurotransmitter role for taurine in the hypothalamus. This was accomplished by comparing the characteristics and properties of the binding, uptake, and release of taurine with those for the classical neurotransmitters which satisfy the criteria for a neurotransmitter. On such a comparative basis, the characteristics of taurine uptake satisfy the neurotransmitter criterion of inactivation of taurine in the hypothalamus. However, the observed characteristics of taurine binding and release in the hypothalamus do not satisfy the respective neurotransmitter criteria of specific receptors and Ca2+-dependent evoked release. Therefore, solely on the basis of the experimental observations reported herein, we must conclude that taurine apparently does not function as a neurotransmitter in the hypothalamus. Two uptake systems were found in the P2 fraction, a high affinity uptake system and a low affinity uptake system. Uptake systems for taurine have previously been reported in glial and nerve cell homogenates, and therefore, because of the known contamination of crude synaptosomal preparations with glial particles, we sought to determine the cellular origin of the two taurine uptake systems in our crude preparation. Using a variety of diverse biochemical techniques such as hypo-osmotic shock, release experiments and Arrhenius plots, we determined that physical changes of the media or depolarizing stimuli which would influence neuronal and glial cell particles differently, also had differing effects on high and low affinity taurine uptake or its release from the respective uptake compartments. We conclude that the high affinity taurine uptake system/compartment is located on/in neuronal membranes/particles/particles and that the low affinity taurine uptake system/compartment is located on/in neuronal membranes/particles and that model for the differential cellular transport and compartmentalization of taurine into neuronal and glial cells has important implications concerning its possible role in the CNS.

Animals↗
PubMed1986

Properties of spontaneous and evoked release of taurine from hypothalamic crude P2 synaptosomal preparations.

Recent studies suggest that taurine may function as a neurotransmitter in the hypothalamus. We examined this role for taurine by characterizing the high K+- and veratridine-evoked release, and the spontaneous release of [3H]taurine from superfused synaptosomal pellets (once-washed crude P2 fractions) prepared from rat hypothalami. Exposure of washed crude P2 synaptosomal pellets which had been preloaded with a concentration of [3H]taurine (1.5 microM) in the high affinity uptake range to either 56 mM K+ or 100 microM veratridine evoked a Ca2+-independent release of [3H]taurine. Exposure of washed crude P2 synaptosomal pellets, which had been preloaded with a concentration of [3H]taurine (2 mM) in the low affinity uptake range to 56 mM K+, induced a Ca2+-independent release of [3H]taurine, whereas 100 microM veratridine did not, either in the presence or absence of Ca2+. These observations support the hypothesis that the high affinity uptake system is located on neuronal membranes with evoked release occurring from a non-vesicular pool of taurine in the neuronal cytoplasm. In contrast, the low affinity uptake system appears to be located on glial membranes with evoked release occurring from a pool of taurine in the glial cytoplasm. Spontaneous [3H]taurine efflux from the high affinity uptake pool in the crude P2 synaptosomal pellet was not Ca2+-dependent. Furthermore, efflux was significantly reduced when NaCl was osmotically replaced with choline chloride in the superfusing medium. These observations suggest that the evoked release of taurine is not simply a reversal of the Na+-dependent high affinity taurine uptake carrier, but accomplished through some other unknown mechanism. The results presented in this report do not support a neurotransmitter role for taurine in the hypothalamus.

Animals↗