[The salt content of food: a public health problem]. / Le sel dans l'alimentation: un problème de santé publique.

Salt abuse in nutrition may exert harmful effects on health, increasing arterial hypertension and its cardiovascular consequences. It is a risk factor, particularly for older subjects and those having chronic diseases such as arterial hypertension, some renal diseases, and obesity. In subjects more particularly vulnerable, the maintenance of sodium balance, which is mainly aldosterone dependent, is perturbed. Although the use of salt for food preservation has greatly declined, it remains a serious risk factor. Excessive salt intake however results more often from poor dietary habits. The WHO and AFSSA have advised to reduce daily salt intake to 5 g, whereas it is currently about 9-10 g. In spite of repeated warnings, salt abuse remains the causal agent for many disease conditions, mainly arterial hypertension. That is why legislative measures should be taken in order to limit the salt content of food industry products, particularly as a preservative in foods. A large-scale public information campaign would be necessary with participation of public health partners, particularly physicians and pharmacists.

[Pharmacology, an innovative factor in therapeutic research]. / La pharmacologie, facteur d'innovation en recherche thérapeutique.

As members of the pharmacology training group set up by the committee of pharmacological science of the French Academy of Pharmacy, we examine the situation of pharmacology in drug discovery. Today, it is obvious that by integrating genome sequencing, cellular and molecular biology, and bioinformatics, pharmacology has become a cross-disciplinary science. Pharmacologists must become knowledgeable in a wide range of domains, using the major points in each to direct them towards the discovery and development of new therapeutic agents. It is also clear that pharmacology remains a major factor in the different steps of drug discovery, from the molecular and cellular stages, to clinical and pharmaceutical developments.

The low molecular weight heparin, enoxaparin, limits infarct size at reperfusion in the dog.

OBJECTIVE: Heparin (HEP) is used in the post-thrombolytic state to prevent vessel reocclusion, thereby aiding myocardial salvage. Side effects limit its benefits, but besides anticoagulant activity HEP has diffuse actions that may be potentially beneficial to jeopardized reperfused myocardium. This study compares the effect of therapeutic doses of HEP and enoxaparin (ENOX), a low molecular weight heparin, and to streptokinase (SK), on infarct size. METHODS: The left anterior descending coronary artery was occluded in dogs for 90 min, followed by 6 h of reperfusion with a residual critical stenosis in place. Five min before reperfusion, HEP (2800 IU) was injected i.v., and perfused at 500 IU/h until sacrifice in group 2, while groups 3 and 4 received ENOX (2128 anti-Xa IU i.v.) followed by 380 anti-Xa IU/h. Group 4 was also given 500,000 IU SK over 30 min before reperfusion beginning at 55 min of occlusion (ENOX + SK), while group 5 received only SK. Controls (CON, group 1) received saline. P-selectin mediated platelet-neutrophil rosettes formation was also tested in vitro in the presence of HEP and ENOX. RESULTS: The area at risk delimited by dye perfusion was statistically similar among groups. Covariance analysis between infarct size (% of area at risk) delimited with triphenyltetrazolium and collateral flow measured with radioactive microspheres confirmed that groups given ENOX (21.6 +/- 5.5%) and ENOX + SK (24.9 +/- 3.9%) developed smaller infarcts (P < 0.05) than CON (48.1 +/- 4.5%), as opposed to HEP (32.2 +/- 3.6%) and SK (46.8 +/- 3.4%) groups. 111In-platelet counts in the infarct were reduced significantly by 64% in the ENOX group as compared to CON, and to a lesser extent (42%, n.s.) in the ENOX + SK group, but were not reduced by HEP and SK treatments. Neutrophil accumulation in the infarcts was decreased significantly and by more than 75% in the ENOX and ENOX + SK groups versus CON, but not in the HEP and SK groups. Also, only ENOX (10-100 micrograms/ml) significantly inhibited platelet-neutrophil rosettes formation in a plasmatic milieu. CONCLUSIONS: The ENOX treatment, as opposed to that of HEP, reduces myocardial platelet and neutrophil accumulations, and limits infarct size when given just before and during reperfusion. The benefits of ENOX on infarct size were not modified by SK, and may be related, at least in part, to an interaction with P-selectin-mediated cell adhesion.

Enoxaparin in experimental stroke: neuroprotection and therapeutic window of opportunity.

BACKGROUND AND PURPOSE: Heparin and heparinoids have long been proposed for stroke treatment. This study investigates the effect of enoxaparin (Lovenox, Clexane), a low-molecular-weight heparin, on functional outcome (neuroscore) and lesion size in stroke models with reversible and irreversible cerebral ischemia using middle cerebral artery occlusion (MCAO) in the rat. METHODS: Ischemia was induced in rats by transient occlusion for 2 hours or by permanent electrocoagulation of the left MCA. Forty-eight hours after ischemia, neurological deficit was evaluated by scoring sensorimotor functions and ischemic damage was quantified by histological evaluation of lesion volumes. RESULTS: After transient MCAO, enoxaparin at 2x1.5 mg/kg IV (2 and 24 hours after insult) significantly reduced lesion size by 30% (P<0.05) and improved neuroscore (P<0.01). This significant effect on lesion size and neuroscore was still evident when treatment was started 5 hours after insult. Administered under the same protocol with a 5 hours delay post permanent MCAO, enoxaparin reduced lesion size by 49% (P<0.05) and improved neuroscore (P<0.01). CONCLUSIONS: This study indicates that standard nonhemorrhagic doses of enoxaparin reduce ischemic damage with a wide therapeutic window. In addition to its anticoagulant properties, other properties of enoxaparin could act in synergy to explain its neuroprotective profile in ischemia. Thus clinical application of enoxaparin treatment in stroke warrants serious consideration.

Binding sites for a peripheral type benzodiazepine antagonist ([3H]PK 11195) in human iris.

Peripheral-type benzodiazepine binding sites have been characterized on sections of 8 normal human iris/ciliary-body preparations. Saturability was determined at 25 degrees C with [3H] PK 11195 (1 nM) a specific ligand of peripheral type sites. The studies revealed a single class of binding sites for PK 11195 with a nanomolar range affinity (KD = 1.45 nM) and a maximal capacity (Bmax) of 35.5 fmol/mg protein. The displacement potency order of the benzodiazepines tested suggest that these sites belong to the peripheral type: PK 11211 (IC50 = 12 nM) greater than Ro 5-4864 (IC50 = 770 nM) greater than clonazepam (IC50 = 20,000 nM). The present data demonstrate that high affinity binding sites for peripheral type benzodiazepines are present in human iris/ciliary-body. This tissue is therefore a suitable tool for evaluation of the putative functional role of these binding sites.

2-Amino-6-trifluoromethoxy benzothiazole, a possible antagonist of excitatory amino acid neurotransmission--I. Anticonvulsant properties.

2-Amino-6-trifluoromethoxy benzothiazole (PK 26124) prevented convulsions induced in rodents by maximal electroshock, inhibitors of the synthesis of gamma-aminobutyric acid (GABA) and ouabain, but was inactive against seizures provoked by GABA antagonists, unlike diazepam, chlordiazepoxide, phenobarbital and valproic acid. 2-Amino-6-trifluoromethoxy benzothiazole prevented seizures induced by sound stimuli in DBA/2 mice (ED50 = 0.66; 2.1 and 4.1 mg/kg, i.p. according to the seizure component), postural seizures in El mice (ED50 = 7.5 mg, i.p.) and seizures induced by photic stimulation in the baboon, Papio papio, at 4 and 8 mg/kg (i.v.). This spectrum of anticonvulsant activity closely resembles that reported previously for dicarboxylic amino acid antagonists. Indeed, PK 26124 prevented seizures induced by L-glutamate (ED50 = 8.5 mg/kg, i.p.) or by kainate (ED50 = 9.25 mg/kg, i.p.) and tremors induced by harmaline (ED50 = 2.5 mg/kg, i.p.) In these tests diazepam was inactive (L-glutamate) or as potent as PK 26124 (kainate, harmaline), whereas it was 10-20 times more potent than PK 26124 against seizures induced by inhibitors of the synthesis of GABA. Together, these data suggest that PK 26124 possesses antagonistic properties of excitatory dicarboxylic amino acids, which may contribute to its anticonvulsant action.

2-Amino-6-trifluoromethoxy benzothiazole, a possible antagonist of excitatory amino acid neurotransmission--II. Biochemical properties.

Two models have been chosen to study the effect of 2-amino-6-trifluoromethoxy benzothiazole (PK 26124) on excitatory amino acid neurotransmission: the pool of cyclic guanosine monophosphate (cGMP) in the cerebellum and the release of acetylcholine in the striatum and olfactory tubercles. The release of acetylcholine induced by N-methyl-DL-aspartate in the striatum and olfactory tubercles was antagonized by PK 26124 which was less potent on the release of acetylcholine induced electrically. The increase in levels of cGMP in the cerebellum induced by excitatory amino acids such as glutamate and quisqualate was antagonized by PK 26124, but the drug was inactive against N-methyl-DL-aspartate, L-aspartate, kainate and cysteine sulphinate. In vivo it antagonized the increases of cGMP in the cerebellum elicited by all these excitatory compounds. All these results are compatible with a possible antagonism by PK 26124 of the excitatory amino acid neurotransmission and may explain its anticonvulsant properties.

The 5-hydroxytryptamine-releasing properties of two epimer quinoline derivatives.

Two epimer quinoline derivatives, PK 5078 and PK 7059, have been shown to be potent at releasing 5-HT from blood platelets. Moreover PK 5078 was also a potent and selective inhibitor of the uptake of 5-HT, being about 20 times as active as clomipramine. Both drugs, like p-chloroamphetamine, released 5-HT but did not inhibit MAO-A. Whilst p-chloroamphetamine seemed to be active on the cytoplasmic pool of 5-HT and reserpine on the vesicular pool, PK 5078 and PK 7059 were effective first on the vesicular pool and then on the cytoplasmic pool. The quinoline derivatives were devoid of the typical side-effects of amphetamine-like drugs, i.e. hyperactivity, anorexia and group toxicity. For these reasons PK 5078 and PK 7059 can be considered to be a new type of selective 5.HT-releasing drug.

Biochemical evidence that 2-phenyl-4[(4-piperidinyl) ethyl]quinoline, a quinoline derivative with pure anticonflict properties, is a partial agonist of benzodiazepine receptors.

The atypical profile of 2-phenyl-4[2-(4-piperidinyl) ethyl]quinoline (PK 8165), a quinoline derivative with pure anticonflict properties, seems to be due to the fact that this compound is a partial agonist of benzodiazepine receptors. The drug PK 8165 is a competitive inhibitor of benzodiazepine binding sites with a Hill coefficient near unity. Opposite to 3-methyl-6-(3-trifluoromethylphenyl)2,4-triazolo(4,5-b)pyridazine (CL 218,872) it was unable to discriminate between BZ1 and BZ2 receptors in sections of brain. However, modulation by gamma-aminobutyric acid (GABA) and the effect of photolabelling by flunitrazepam on the affinity of PK 8165 indicated that GABA or photolabelling shifts of PK 8165 were between full agonists and antagonists. By itself PK 8165 was unable to modify the levels of cGMP in the cerebellum, but potentiated the lowering of levels of cGMP by diazepam and did not present antagonistic properties of this effect.

3-(4-Piperidinylalkyl)indoles, selective inhibitors of neuronal 5-hydroxytryptamine uptake.

A series of 3-(4-piperidinylalkyl)indoles was synthesized and tested as uptake inhibitors of biogenic amines. Some of these compounds are potent and very selective in blocking the 5-hydroxytryptamine (5-HT) uptake, as evidenced by biochemical data and behavioral tests. A discussion on structure-activity relationships is given. The most interesting member of the series, indalpine, 3-[2-(4-piperidinyl)ethyl]indole (1), was selected for clinical studies.

4-Amino-6-chloro-2-piperazinopyrimidines with selective affinity for alpha 2-adrenoceptors.

A series of 4-amino-6-chloro-2-piperazinopyrimidines were synthesized and evaluated for their ability to interact with alpha 1- and alpha 2-adrenoceptors in vitro in binding assays using [3H]WB-4101, [3H]clonidine, and [3H]idazoxan as radioligands. Some compounds were also tested as inhibitors of [3H]spiroperidol binding. Several members of this series showed high and selective affinity for alpha 2-adrenoceptors. The nature of the 4-amino substituent seems to be the most critical factor in determining the potency at these receptors.

2-Amino-6-chloro-4-(N-methylpiperazino)pyrimidines, inhibitors of spiroperidol binding.

A series of 30 6-chloro-2,4-diaminopyrimidines was synthesized and tested in vitro as inhibitors of [3H]spiroperidol binding. The affinity for the dopamine receptor was shown to be related to the 6-chloro-4-(N-methyl-piperazine)pyrimidine structure bearing a NH2 or NHR1 group as a substituent in position 2, provided that R1 was not an alpha branched alkyl group. The nature of the substituent in position 5 is also of importance for the affinity; 2-(benzylamino)-6-chloro-4-(N-methylpiperazino)-5-(methylthio)pyrimidine (22) is the most active member of the series. Molecular structures of three compounds were analyzed by X-ray diffraction and PCILO computation.

On the relationship between molecular mass and anticoagulant activity in a low molecular weight heparin (enoxaparin).

A low molecular weight heparin (enoxaparin, mean molecular weight approximately 4,400) was separated by gel chromatography into eight different fractions with a narrow distribution around the following mean molecular weights: 1,800, 2,400, 2,900, 4,200, 6,200, 8,600, 9,800 and 11,000. We compared the influence of enoxaparin on the generation of thrombin in plasma to that of the eight fractions. We determined: a) the % of material with high affinity to antithrombin III (HAM) and the % of HAM above the critical chain length necessary to allow for thrombin inhibition (ACLM), b) the specific catalytic activity on the decay of endogenous thrombin, and c) the inhibition of over-all thrombin formation in the extrinsic and the intrinsic pathway. From b and c we calculated the inhibition of prothrombin conversion in these pathways. We found that a) there is a gradual decrease of the HAM fraction with decreasing molecular weight; b) the specific catalytic activity for the inactivation of thrombin does not vary significantly between the fractions when expressed in terms of ACLM; c) the potency to inhibit prothrombin conversion does not vary significantly between the fractions when expressed in terms of HAM.

Inhibition of platelet adhesion to fibrin(ogen) in flowing whole blood by Arg-Gly-Asp and fibrinogen gamma-chain carboxy terminal peptides.

We have employed synthetic peptides with sequences corresponding to the integrin receptor-recognition regions of fibrinogen as inhibitors of platelet aggregation and adhesion to fibrinogen- and fibrin-coated surfaces in flowing whole blood, using a rectangular perfusion chamber at wall shear rates of 300 s-1 and 1,300 s-1. D-RGDW caused substantial inhibition of platelet aggregation and adhesion to fibrinogen and fibrin at both shear rates, although it was least effective at blocking platelet adhesion to fibrin at 300 s-1. RGDS was a weaker inhibitor, and produced a biphasic dose-response curve; SDRG was inactive. HHLGGAKQAGDV partially inhibited platelet aggregation and adhesion to fibrin(ogen) at both shear rates. These results support the identification of an RGD-specific receptor, most likely the platelet integrin glycoprotein IIb:IIIa, as the primary receptor responsible for platelet:fibrin(ogen) adhesive interactions under flow conditions, and indicate that platelet adhesion to surface bound fibrin(ogen) is stabilized by multivalent receptor-ligand contacts.

Binding in vitro of pipequaline (45319 RP) onto plasma proteins and blood cells in man.

Serum binding of pipequaline, a new anxiolytic drug, was studied in vitro by equilibrium dialysis. The percent binding in serum is high, 96.3%, and remains constant within the range of therapeutic concentrations. Investigations performed on isolated proteins with a wide range of concentrations showed one site with a high affinity constant (Ka = 450,000 M-1) for alpha 1-acid glycoprotein and two sites with a lower affinity constant (Ka = 58,000 M-1) for human serum albumin. Binding to lipoproteins was saturable, with an affinity constant of 22,000 less than or equal to Ka less than or equal to 35,000 M-1. Over the range of therapeutic concentrations, the ratio of pipequaline concentrations in serum and red blood cells remained constant (14.4%) and was shown to be dependent on the free fraction of pipequaline in serum.

Characterization of solubilized "peripheral type" benzodiazepine binding sites from rat adrenals by using [3H]PK 11195, an isoquinoline carboxamide derivative.

"Peripheral type" benzodiazepine binding sites have been solubilized with digitonin. Binding site density for the solubilized material is increased 1.7 times compared to membranes. A decrease in the affinity for [3H]-PK 11195 (a new ligand for the peripheral type benzodiazepine binding sites) was also observed. Pharmacological specificity of displacing agents was conserved during solubilization. The apparent molecular weight determined by gel filtration was 215,000 +/- 20,000. The high Bmax value of the solubilized preparation (greater than 50 pmole/mg protein) makes it advantageous as the starting point for a purification procedure.

Characterization of peripheral type benzodiazepine binding sites in human and rat platelets by using [3H]PK 11195. Studies in hypertensive patients.

Peripheral type benzodiazepine binding sites have been studied in human and rat platelets and platelet membranes by using PK 11195 (1-(2-chlorophenyl)-N-methyl-N-(1-methyl propyl)-3-isoquinolinecarboxamide) as a ligand. [3H]PK 11195 binding to the intact cells and membranes is saturable, with a high affinity and presents the pharmacological specificity corresponding to the peripheral binding sites (PK 11195 greater than RO5-4864 greater than diazepam greater than clonazepam). [3H]PK 11195 affinity is not affected by cell lysis, but there is a loss of binding capacity, contrarily to RO5-4864 whose affinity is greatly diminished. For this reason [3H]RO5-4864 binding can only be demonstrated in intact cells. Furthermore opposite to RO5-4864, PK 11195 affinity is not decreased by increasing temperatures. No difference was found between binding parameters (KD and Bmax) for [3H]PK 11195 between normotensive and hypertensive subjects. The very high binding capacity of human and rat platelets (Bmax greater than pmole/10(8) cells) makes them a good biological model for studying the physiological significance of "peripheral type" benzodiazepine binding sites.

Detection of binding sites for spiroperidol on leukemic cells: its value for the phenotype characterization of lymphoid leukemias.

The specific binding of the dopamine antagonist spiroperidol was studied in leukemic cell samples of various phenotypes. Among these only B-cell samples from chronic lymphocytic leukemias (7/7) and some "null" cell samples from acute lymphoblastic leukemias (2/7) showed specific binding. B cells from a prolymphocytic leukemia were negative as were also T-lymphoïd and non-lymphoïd leukemic cells at different stages of maturation. This pattern can be clearly correlated with the previous results obtained with normal blood cells and on cell lines. Moreover, it suggests that the detection of spiroperidol binding sites could provide a new means of distinguishing different phenotypes among B cells and early lymphoïd cells. Our results open the way to further studies which might show a correlation between spiroperidol binding sites and the new immunological markers defining subsets among non-T lymphoïd cells, as well as defining their physiological meaning.

Enoxaparin reduces brain edema, cerebral lesions, and improves motor and cognitive impairments induced by a traumatic brain injury in rats.

Traumatic brain injury (TBI) is often accompanied by secondary ischemia due, in part, to edema-induced blood vessel compression. Enoxaparin, a low-molecular weight heparin, which is efficacious in models of myocardial and brain ischemia was studied in lateral fluid percussion-induced TBI in rats. Enoxaparin was administered 2 h post-TBI at 0.5 mg/kg i.v. followed by 4 x 0.5, 4 x 1, or 4 x 2 mg/kg s.c. over 30 h. Brain edema was measured in the hippocampus, temporal cortex and parietal cortex. Edema was reduced by enoxaparin (0.5 + 4 x 0.5 mg/kg) in the hippocampus (-53%, p = 0.07) and the parietal cortex (-39%, ns). At 0.5 + 4 x 1 mg/kg edema was reduced in the hippocampus (-63%, p < 0.05) and the parietal cortex (-47%, p = 0.06). At 0.5 + 4 x 2 mg/kg, the reduction was more important in the hippocampus (-69%, p < 0.01) and in the parietal cortex (-50%, p < 0.05). No reduction was seen in the temporal cortex. The lesion size was reduced by enoxaparin at 0.5 + 4 x 1 mg/kg (-50%, p < 0.05), and at 0.5 + 4 x 2 mg/kg (-35%, ns). The neurological deficit evaluated with a 9-point scale was also improved with enoxaparin at 0.5 + 4 x 1 mg/kg 1 week post-TBI (p < 0.05). The cognitive impairment evaluated with a Lashley maze task was improved with enoxaparin (0.5 + 4 x 1 mg/kg) from 48 h (p < 0.05) to 2 weeks post-TBI (p < 0.01). Our results demonstrate for the first time that enoxaparin significantly reduces the brain contusion and edema, and improves the functional outcomes induced by a TBI. Therefore, enoxaparin could be a candidate drug to treat acute brain-injured patients.

Central dopaminergic neurons during development of genetic and DOCA-salt hypertension in the rat.

The in vivo binding of [3H]spiroperidol was measured in discrete areas of the brain in 7-, 9- and 16-week-old spontaneously hypertensive rats (SHR) and age-matched normotensive Wistar-Kyoto (WKY) controls. An increase in the [3H]spiroperidol binding in the striatum, tuberculum olfactorium and frontal cortex but not in the cerebellum was detected at all ages in SHR. The increase was more pronounced in 7- than in 9- or 16-week-old SHR. In vitro data indicated an increase in Bmax but no variation in Kd in the striatum of 7-week-old SHR. Moreover no difference was detectable in the dopaminergic cell bodies (A9, A10). This increase was specific to [3H]spiroperidol binding sites since no difference was observed in the in vivo binding of [3H]QNB and [3H]LSD in the same brain regions. No variation in dopamine level or dopamine utilization, as estimated by measuring the disappearance of the amine induced by alpha-methyl-p-tyrosine, was observed. The DOPA accumulation after injection of the DOPA decarboxylase inhibitor NSD 1015 was greater in the tuberculum olfactorium from 7-week-old SHR. An increase in [3H]spiroperidol binding sites was also observed in the striatum and tuberculum olfactorium after 7 weeks of DOCA-salt treatment. These results suggest that dopaminergic neurons might be implicated in the onset of hypertension in the rat.