Biomedical subjects

H Rochat

Publications and source records attributed to H Rochat.

At least 55 records · Page 3Linked to original sources

PubMed1998

Adenosine and the nervous system: pharmacological data and therapeutic perspectives.

1. Adenosine acts on a family of G-protein-coupled receptors called purinoreceptors. 2. Four subtypes have been cloned and pharmacologically characterized. 3. The principal pharmacological data and structure-function relations for agonist interactions with P1 receptors are presented. 4. We conclude that the potent role of adenosine in the nervous system may be interesting for the development of drugs targeted at purines and their receptors.

Adenosine↗

PubMed1998

Voltage sensor-trapping: enhanced activation of sodium channels by beta-scorpion toxin bound to the S3-S4 loop in domain II.

Polypeptide neurotoxins alter ion channel gating by binding to extracellular receptor sites, even though the voltage sensors are in their S4 transmembrane segments. By analysis of sodium channel chimeras, a beta-scorpion toxin is shown here to negatively shift voltage dependence of activation and enhance closed state inactivation by binding to a receptor site that requires glycine 845 (Gly-845) in the S3-S4 loop at the extracellular end of the S4 segment in domain II of the alpha subunit. Toxin action requires prior depolarization to drive the S4 voltage sensors outward, but these effects are lost in the mutant G845N. The results reveal a voltage sensor-trapping model of toxin action in which the IIS4 voltage sensor is trapped in its outward, activated position by toxin binding.

Amino Acid Sequence↗

PubMed1998

Adenosine and migraine.

BACKGROUND: Adenosine is a powerful natural vasodilator that participates in the control of cerebral and meningeal blood flow. In this context, it could be involved in the pathophysiology of migraine, since it was previously reported that intravenous adenosine can precipitate crises in migraine patients. METHODS: We have investigated circulating adenosine levels in 12 patients suffering from migraine without aura, during crises and in crisis-free periods, and have compared the levels noted to those of a population of 10 controls. To determine if there are interactions between adenosine and serotonin, we examined the effect of adenosine and antagonists on the uptake and the release of (14C) serotonin by platelets. RESULTS AND CONCLUSION: We have reached a dual conclusion: 1) during migraine headaches there is an increase (mean 68%) in circulating adenosine levels and this increase may participate in cephalalgia; 2) activation of A2 receptors by adenosine causes a dose-dependent serotonin uptake by platelets. This inhibition of uptake could participate in the rapid elimination of serotonin in migraine sufferers. As a result of this, the use of adenosine antagonists could be an effective complementary treatment for migraine.

Adenosine↗

PubMed1998

Interactions between cyclosporine A and adenosine in kidney transplant recipients.

Adenosine is involved in a large number of physiological processes including immune response and vasomotor function. But its precise involvement in renal physiology is poorly understood. We have investigated the putative relationships between cyclosporine A (CsA) and adenosine (ADO) metabolism in kidney transplant recipients (KTR). We first compared ADO plasma levels in three groups of patients and in 10 controls: the first group (N = 14) was composed of CsA-treated KTR; the second group (N = 5) was KTR not treated with CsA, and the third (N = 6) was chronic kidney failure patients. We also measured ADO plasma level in two KTR treated with FK506, a CsA analog. ADO plasma levels in CsA-treated KTR were significantly higher (mean 0.76 microM +/- 0.27) than in the control group (mean 0.31 +/- 0.13; Mean-Whitney test, S = 8.5; P = 2.1 x 10(-4)) and than in the chronic kidney failure group (0.37 +/- 0.16, Mann-Whitney, S = 5.5; P = 1.6 x 10(-3)). In CsA-treated KTR, CsA and ADO plasma levels were significantly correlated (Spearman's, r = 0.8, P = 1.9 x 10(-3)). No significant differences in ADO plasma levels were found between patients with chronic kidney failure and controls (P < 0.05). ADO plasma levels in KTR not treated with CsA were in the same range as those in controls. Finally, the ADO plasma level was increased in the two FK506-treated patients. We also investigated the action of CsA on ADO plasma degradation and uptake by erythrocytes in vitro. No interaction between adenosine deaminase and CsA was found because CsA, in the presence of adenosine deaminase, did not modify the plasma half-life of ADO. Conversely, in the presence of CsA (500 and 1000 ng/ml), the uptake of ADO by erythrocytes was significantly decreased in adenosine deaminase-free samples (analysis of variance, P = 1.8.10(-3) and 1.2 x 10(-4), respectively). We conclude that ADO plasma levels are significantly elevated and correlate with CsA blood level in CsA-treated KTR, and that these high levels are due to CsA inhibition of ADO uptake by red cells. Since ADO and metabolites have well known immunosuppressive and vascular effects, ADO is likely to participate in the immune defect and in the vasoconstriction induced by CsA.

Adenosine↗

PubMed1998

The hormonal response to stress is not modified by the dramatic decrease in prolactin plasma concentration during surgery for microprolactinoma.

OBJECTIVES: To determine the endocrine response to surgical stress in a homogeneous population of 36 women with microprolactinomas, particularly to evaluate the effect of the sharp decrease in plasma prolactin on stress induced hormonal secretion. In addition, the effects of exogenous opiates on prolactin secretion were studied. METHODS: The plasma kinetics of cortisol, prolactin, ACTH, GH, and beta-endorphin like immunoreactivity (beta-ELI) were analysed by including patients operated on with strict anaesthetic and surgical protocols, and by sampling blood every 10 minutes, starting at premedication up to 3 hours after induction. RESULTS AND CONCLUSIONS: (a) Surgical stress or opiate administration did not induce prolactin release in patients with microprolactinoma. (b) The dramatic decrease in prolactin concentrations have apparently no effect on the release of other hormones involved in stress. (c) The existence of an early GH peak, independently of any surgical procedure, strongly suggests that GH is released by surgical stress whereas beta-endorphin is secreted in response to pain. Thus GH may be a useful marker of surgical stress.

Adolescent↗

PubMed1997

Influence of a NH2-terminal extension on the activity of KTX2, a K+ channel blocker purified from Androctonus australis scorpion venom.

A cDNA encoding a short polypeptide blocker of K+ channels, kaliotoxin 2 (KTX2), from the venom of the North African scorpion Androctonus australis was expressed in the periplasmic space of Escherichia coli. KTX2 was produced as a fusion protein with the maltose binding protein followed by the recognition site for factor Xa or enterokinase preceding the first amino acid residue of the toxin. The fully refolded recombinant KTX2 (rKTX2) was obtained (0.15-0.30 mg/l of culture) and was indistinguishable from the native toxin according to chemical and biological criteria. An N-extended analogue of KTX2 exhibiting three additional residues was also expressed. This analogue had 1000-fold less affinity for the 125I-kaliotoxin binding site on rat brain synaptosomes than KTX2. Conformational models of KTX2 and its mutant were designed by amino acid replacement using the structure of agitoxin 2 from Leiurus quinquestriatus as template, to try to understand the decrease in affinity for the receptor.

ATP-Binding Cassette Transporters↗

PubMed1997

Monoclonal antibodies neutralizing the toxin II from Androctonus australis hector scorpion venom: usefulness of a synthetic, non-toxic analog.

Scorpion venom contains toxins that act on ion channels. Some are responsible for the noxious effects observed when people are stung by scorpions. The study of the neutralization of these molecules and the production of monoclonal antibodies (mAbs) should prove valuable. Toxin II from Androctonus australis hector scorpion (AahII) is one of the most potent toxins and has been well-characterized and studied. Producing mAbs against such molecules is often difficult due to their toxicity. We used a synthetic, non-toxic analog, (Abu)8-AahII, to obtain mAbs which recognize and neutralize the native toxin AahII. Sets of peptides spanning the entire sequence of AahII were assayed to identify the binding sites of the mAbs. The various mAbs recognized only the largest peptides (12-17 residues). They recognized peptides corresponding to different parts of the AahII sequence, suggesting that several regions of the (Abu)8-AahII sequence mimic AahII epitopes and then elicit mAbs directed against toxin.

Amino Acid Sequence↗

PubMed1997

1H-NMR-derived secondary structure and overall fold of a natural anatoxin from the scorpion Androctonus australis hector.

The venom of the scorpion Androctonus australis hector contains several protein neurotoxins of which structure and structure/activity relationships have been extensively studied. It also contains polypeptides such as Aah STR1, which are not toxic, while having highly similar sequences to fully active toxins. We have determined the solution structure of Aah STR1 by use of conventional two-dimensional NMR techniques followed by distance-geometry and energy minimization. We have demonstrated that, despite its lack of toxicity, Aah STR1 is structurally highly related to anti-mammal scorpion toxins specific for Na+ channels. The calculated structure is composed of a short alpha-helix (residues 26-33) connected by a tight turn to a three-stranded antiparallel beta-sheet (sequences 3-6, 38-41 and 44-48). This beta-sheet is right-handed twisted as usual for such secondary structures. The beta-turn connecting the strands 38-41 and 44-48 belongs to type II'. The overall fold of Aah STR1 is typical of beta-type scorpion toxins. This is, however, the first example of such a fold in Old World scorpion toxins. Either the absence of a basic residue in position 63 or the high mobility of loops, compared to active beta-type neurotoxins, may explain the lack of activity of this protein.

Amino Acid Sequence↗

PubMed1997

Maurotoxin, a four disulfide bridge toxin from Scorpio maurus venom: purification, structure and action on potassium channels.

A new toxin acting on K+ channels, maurotoxin (MTX), has been purified to homogeneity from the venom of the chactoid scorpion Scorpio maurus. MTX is a basic single chain 34 amino acid residue polypeptide, amidated at its C terminal, and crosslinked by four disulfide bridges. It shows 29-68% sequence identity with other K+ channel toxins, and presents an original disulfide pattern, the last two half-cystine residues (31-34) being connected. Although the first three disulfide bonds have not been defined experimentally, modelling based on the structure of charybdotoxin favored two combinations out of six, one of which has two bridges (3-24 and 9-29) in common with the general motif of scorpion toxins. The last bridge would connect residues 13 and 19. MTX inhibits the binding to rat brain synaptosomal membranes of both [125I]apamin, a SK(Ca) channel blocker (IC50 5 nM), and [125I]kaliotoxin, a Kv channel blocker (IC50 30 pM). MTX blocks the Kv1.1, Kv1.2 and Kv1.3 currents expressed in Xenopus oocytes with IC50 of 45, 0.8 and 180 nM, respectively. MTX represents a member of a new class of short toxins with 4 disulfide bridges, active on voltage-dependent K+ channel and also competing with apamin for binding to its receptor.

Amino Acid Sequence↗

PubMed1997

Bot IT2: a new scorpion toxin to study receptor site on insect sodium channels.

The insect-specific Bothus occitanus tunetanus IT2 toxin is distinguishable from other scorpion toxins by its amino acid sequence and effects on sodium conductance. The present study reveals that Bot IT2 possesses in cockroach neuronal membranes a single class of high affinity (Kd = 0.3 +/- 0.1 nM) and low capacity (Bmax = 2.4 +/- 0.5 pmol/mg) binding sites. Competitive binding experiments with several known sodium channel neurotoxins reveal that the Bot IT2 binding site is in close proximity to the other toxins.

Animals↗

PubMed1997

Biochemical and pharmacological characterization of a depressant insect toxin from the venom of the scorpion Buthacus arenicola.

A depressant toxin active on insects, Buthacus arenicola IT2, was isolated from the venom of the North African scorpion B. arenicola and its structural and pharmacological properties were investigated. B. arenicola IT2 is a single polypeptide of 61 amino acid residues, including 8 half-cystines but no methionine and histidine, with a molecular mass of 6835 Da. Its amino acid sequence is 79-95% identical to other depressant toxins from scorpions. When injected into the cockroach Blatella germanica, B. arenicola IT2 induced a slow depressant flaccid paralysis with a LD50 of 175 ng. B. arenicola IT2 has two non-interacting binding sites in cockroach neuronal membranes: one of high affinity (Kd1 = 0.11 +/- 0.04 nM) and low capacity (Bmax1 = 2.2 +/- 0.6 pmol/mg), and one of low affinity (Kd2 = 24 +/- 7 nM) and high capacity (Bmax2 = 226 +/- 92 pmol/mg). Its binding to these two sites was completely inhibited by Leiurus quinquestriatus quinquestriatus IT2, a depressant toxin from L. quinquestriatus quinquestriatus. Reciprocal-binding experiments between B. arenicola IT2 and the excitatory insect-toxin A. australis Hector IT revealed competition between the two toxins for the high-affinity sites of B. arenicola IT2. B. arenicola IT2 has a higher affinity than L. quinquestriatus hebraeus IT2, a depressant toxin from L. quinquestriatus hebraeus. Thus, B. arenicola IT2 represents an interesting tool to study the receptor site for depressant toxins on insect sodium channels.

Amino Acid Sequence↗

PubMed1997

Refined solution structure of the anti-mammal and anti-insect LqqIII scorpion toxin: comparison with other scorpion toxins.

The solution structure of the anti-mammal and anti-insect LqqIII toxin from the scorpion Leiurus quinquestriatus quinquestriatus was refined and compared with other long-chain scorpion toxins. This structure, determined by 1H-NMR and molecular modeling, involves an alpha-helix (18-29) linked to a three-stranded beta-sheet (2-6, 33-39, and 43-51) by two disulfide bridges. The average RMSD between the 15 best structures and the mean structure is 0.71 A for C alpha atoms. Comparison between LqqIII, the potent anti-mammal AaHII, and the weakly active variant-3 toxins revealed that the LqqIII three-dimensional structure is closer to that of AaHII than to the variant-3 structure. Moreover, striking analogies were observed between the electrostatic and hydrophobic potentials of LqqIII and AaHII. Several residues are well conserved in long-chain scorpion toxin sequences and seem to be important in protein structure stability and function. Some of them are involved in the CS alpha beta (Cysteine Stabilized alpha-helix beta-sheet) motif. A comparison between the sequences of the RII rat brain and the Drosophila extracellular loops forming scorpion toxin binding-sites of Na+ channels displays differences in the subsites interacting with anti-mammal or anti-insect toxins. This suggests that hydrophobic as well as electrostatic interactions are essential for the binding and specificity of long-chain scorpion toxins.

Amino Acid Sequence↗

PubMed1997

Purification, structure and activity of three insect toxins from Buthus occitanus tunetanus venom.

One contractive and two depressant toxins active on insect were purified by high-performance liquid chromatography from the venom of Buthus occitanus tunetanus (Bot). The two depressant toxins, BotIT4 and BotIT5, differ only at position 6 (Arg for Lys) and are equally toxic to insects (LD50 to Blatella germanica = 110 ng/100 mg body weight). They show a strong antigenic cross-reaction with a depressive toxin from Leiurus quinquestriatus quinquestriatus (LqqIT2). The two toxins are able to inhibit with high affinity (K0.5 between 2 and 3 nM) the specific binding of the radioiodinated excitatory insect toxin (125I-AaHIT) on its receptor site on Periplaneta americana synaptosomal membranes. These toxins depolarize the cockroach axon, irreversibly block the action potential, and slow down and very progressively block the transmembrane transient Na+ current. The contracturant toxin BotIT1 is highly toxic to B. germanica (LD50 = 60 ng/ 100 mg body weight) and barely toxic to mice (LD50 = 1 microgram/20 g body weight) when injected intracerebroventricularly. It does not compete with 125I-AaHIT for its receptor site on P. americana synaptosomal membranes. On cockroach axon, BotIT1 develops plateau potentials and slows down the inactivation mechanism of the Na+ channels. Thus, BotIT1 belongs to the group of alpha insect-selective toxins and shows a strong sequence identity (> 90%) with Lqh alpha IT and LqqIII, two insect alpha-toxins previously purified from the venom of L. q. hebraeus and L. q. quinquestriatus. respectively.

Action Potentials↗

PubMed1997

In vivo protection against Androctonus australis hector scorpion toxin and venom by immunization with a synthetic analog of toxin II.

A synthetic peptide mimicking the North African scorpion Androctonus australis hector toxin II was designed and produced by chemical solid-phase synthesis. It contains the entire sequence of toxin II (64 amino acid residues), with each half-cystine being replaced by the isosteric residue a-aminobutyric acid, and was thus devoid of disulfide bridges. This construct was totally nontoxic in mice even if large amounts, equivalent to 1000 times the LD50 of the original toxin, were injected by the intracerebroventricular route. The synthetic peptide, either as a monomer or polymerized by means of glutaraldehyde, induced the production of antitoxin neutralizing antibodies in immunized mice and rabbits. After three injections with either the monomeric or polymerized synthetic peptide, the immunized mice were protected against several lethal doses of the corresponding native toxin or scorpion venom. Six months after immunization, the mice were completely protected against challenge with eight LD50 of the original toxin. The protection was better when the polymerized synthetic peptide was used. One month after the start of the immunization program, it showed a good correlation between antibody titer and protection. However, antibody titer decreased with time but protection remained high. This suggests that additional factors other than circulating antibodies play a role in protective activity.

Animals↗

PubMed1997

Toxin III from Leiurus quinquestriatus quinquestriatus: a specific probe for receptor site 3 on insect sodium channels.

Scorpion toxin Lqq III binds to a single class of high affinity (Kd = 72 +/- 19 pM) and low capacity (Bmax = 2.5 +/- 0.2 pmol/mg) binding sites in cockroach neuronal membranes. Its binding was inhibited by Lqh alpha IT (IC50 = 80 +/- 30 pM) and sea-anemone toxin ATX II (IC50 = 2.5 +/- 0.3 nM), suggesting that Lqq III is a specific probe for receptor site 3 on cockroach sodium channels. This was confirmed by competitive binding experiments between 125I-Lqq III and scorpion alpha-toxins which have less toxicity in insects.

Amino Acid Sequence↗

PubMed1997

Plasma fibronectin: predictive factor in gestational hypertension?

Our purpose was to evaluate fibronectin as a marker of endothelial cell injury, and as a test for predicting preeclampsia. A retrospective study was performed from November 1993 to March 1995. Results from 142 women were examined: 108 normal pregnant women and 34 pregnant women with evidence of preeclampsia. The plasma fibronectin concentration was significantly higher in pre-eclamptic gravidas (620 +/- 210 mg/l) than in normotensive gravidae women (390 +/- 130 mg/l). A fibronectin concentration lower than 400 mg/l predicted the non-development of a hypertension with a negative predictive value of 96%. The present findings suggest that fibronectin is rather an exclusion parameter than predictive test for hypertension disorders of pregnancy.

Adult↗

PubMed1996

Chemical synthesis and characterization of maurotoxin, a short scorpion toxin with four disulfide bridges that acts on K+ channels.

Maurotoxin is a toxin isolated from the venom of the Tunisian chactoid scorpion Scorpio maurus. It is a 34-amino-acid peptide cross-linked by four disulfide bridges. Maurotoxin competes with radiolabeled apamin and kaliotoxin for binding to rat-brain synaptosomes. Due to its very low concentration in venom (0.6% of the proteins), maurotoxin was chemically synthesized by means of an optimized solid-phase technique. The synthetic maurotoxin was characterized. It was lethal to mice following intracerebroventricular injection (LD50, 80 ng/mouse). The synthetic maurotoxin competed with 125I-apamin and 125I-kaliotoxin for binding to rat-brain synaptosomes with half-maximal effects at concentrations of 5 nM and 0.2 nM, respectively. Synthetic maurotoxin was tested on K+ channels and was found to block the Kv1.1, Kv1.2, and Kv1.3 currents with half-maximal blockage (IC50) at 37, 0.8 and 150 nM, respectively. Thus, maurotoxin is a scorpion toxin with four disulfide bridges that acts on K+ channels. The half-cystine pairings of synthetic maurotoxin were identified by enzymatic cleavage. The pairings were Cys3-Cys24, Cys9-Cys29, Cys13-Cys19 and Cys31-Cys34. This disulfide organization is unique among known scorpion toxins. The physicochemical and pharmacological properties of synthetic maurotoxin were indistinguishable from those of natural maurotoxin, which suggests that natural maurotoxin adopts the same half-cystine pairing pattern. The conformation of synthetic maurotoxin was investigated by means of circular dichroism spectroscopy and molecular modeling. In spite of its unusual half-cystine pairings, the synthetic-maurotoxin conformation appears to be similar to that of other short scorpion toxins.

Amino Acid Sequence↗

PubMed1996

A new scorpion venom toxin paralytic to insects that affects Na+ channel activation. Purification, structure, antigenicity and mode of action.

A new toxin, BotIT2, with a unique mode of action on the isolated giant axon of the cockroach Periplaneta americana and DUM (dorsal unpaired median) neurons, has been purified from the venom of the scorpion Buthus occitanus tunetanus. Its structural, antigenic and pharmacological properties are compared to those of three other groups of neurotoxins found in Buthidae scorpion venoms. Like excitatory, depressant and alpha-type insect-selective neurotoxins, BotIT2 is toxic to insects, but shows the following common and distinctive characteristics. (a) As alpha-type toxins, BotIT2 lack strict selectivity to insects; they have measurable but low toxicity to mice. (b) As depressant toxins and unlike alpha-type toxins, BotIT2 is able to displace iodinated AaHIT from its binding sites in insect neuronal membranes. This indicates that the binding site for BotIT2 is identical, contiguous or in allosteric interaction with that of AaHIT and depressant toxins. (c) The BotIT2 amino acid sequence shows strong similarity to depressant toxins. However, unexpectedly, despite this high sequence similarity, BotIT2 shares moderate cross-antigenic reactivity with depressant toxins. (d) Voltage and current-clamp studies show that BotIT2 induces limited depolarization concomitantly with the development of depolarizing after potential, repetitive activity and later plateau potentials terminated by bursts. Under voltage-clamp conditions, BotIT2 specifically acts on Na+ channels by decreasing the peak Na+ current and by simultaneously inducing a new current with very slow activation/deactivation kinetics. The voltage dependence of this slow current is not significantly different from that of the control current. These observations indicate that BotIT2 chiefly modifies the kinetics of axonal and DUM neuronal membrane Na(+)-channel activation.

Amino Acid Sequence↗