Hyperosmolar D-mannitol reverses the increased membrane excitability and the nodal swelling caused by Caribbean ciguatoxin-1 in single frog myelinated axons

The effects of hyperosmolar D-mannitol were studied on single frog myelinated nerve fibres previously poisoned with Caribbean ciguatoxin-1 (C-CTX-1), a new toxin isolated from the pelagic fish Caranx latus inhabiting the Caribbean region. In current-clamped myelinated axons, C-CTX-1 (50-120 nM) caused spontaneous and repetitive action potential discharges after a short delay. In addition, the toxin produced a marked swelling of nodes of Ranvier of myelinated axons that reached a steady state within about 90 min, as revealed by using confocal laser scanning microscopy. The increased excitability and the nodal swelling caused by C-CTX-1 were prevented or reversed by an external hyperosmotic solution containing 100 mM D-mannitol. Moreover, the C-CTX-1-induced nodal swelling was completely prevented by the blockade of voltage-sensitive sodium channels by tetrodotoxin (TTX). It is suggested that C-CTX-1, by increasing nerve membrane excitability, enhances Na(+) entry into nodes of Ranvier through TTX-sensitive sodium channels, which directly or indirectly disturb the osmotic equilibrium between intra- and extra-axonal media resulting in an influx of water that was responsible for the long-lasting nodal swelling. The fact, that hyperosmolar D-mannitol either reversed or prevented the neurocellular actions of C-CTX-1, is of particular interest since it provides the rational basis for its use to treat the neurological symptoms of ciguatera fish poisoning in the Caribbean area.     

Orthostatic intolerance in the chronic fatigue syndrome

This study aims to investigate the prevalence and pathophysiology of orthostatic intolerance (OI) and its potential contribution to symptoms of a group of unselected patients with chronic fatigue syndrome (CFS). Seventy five patients (65 women, 10 men) with CFS were evaluated. During an initial visit, a clinical suspicion as to the likelihood of observing laboratory evidence of OI was assigned. Laboratory investigation consisted of beat-to-beat recordings of heart rate, blood pressure (Finapres), and stroke volume (impedance cardiograph) while supine and during 80 degrees head-up tilt (HUT), during rhythmic deep breathing (6 breaths/min) and during the Valsalva maneuver. The responses of 48 age-matched healthy controls who had no history of OI were used to define the range of normal responses to these three maneuvers. Forty percent of patients with CFS had OI during head-up tilt. Sixteen exhibited neurally-mediated syncope alone, seven tachycardia (> 35 bpm averaged over the whole of the head-up tilt) and six a mixture of tachycardia and syncope. Eight of 48 controls exhibited neurally-mediated syncope. The responses to the Valsalva maneuver and to deep breathing were similar in controls and patients. On average, the duration of disease and patient age were significantly less and the onset of symptoms was more often subacute in patients with OI than in those without OI. We conclude that there exists a clinically identifiable subgroup of patients with CFS and OI that differs from control subjects and from those with CFS without OI for whom treatment specifically aimed at improving orthostatic tolerance may be indicated.     

Reduction in susceptibility to Brugia malayi of F2 progeny of Aedes togoi treated with ethyl methanesulfonate

The susceptibility to Brugia malayi infection was tested in F2 female progeny derived from male and female Aedes togoi treated with ethyl methanesulfonate (EMS). Three-day-old males and females were treated with 0.025, 0.050, and 0.075, 0.10, 0.15, or 0.20% EMS by allowing them to feed for 5 days on sugar cubes containing EMS and then mated at random. Percentage of susceptibility and mean number of infective larvae (L3) in F2 females were analyzed over a 2-wk period. Reductions in susceptibility were significant in the F2 populations arising from the 3 highest EMS concentrations. F2 infections were reduced by 80%, indicating that EMS-induced mutations affect loci associated with filarial nematode susceptibility.     

Human and rat liver UDP-glucuronosyltransferases are targets of ketoprofen acylglucuronide.

Acylglucuronides formed from carboxylic acids by UDP-glucuronosyltransferases (UGTs) are electrophilic metabolites able to covalently bind proteins. In this study, we demonstrate the reactivity of the acylglucuronide from the nonsteroidal anti-inflammatory drug, ketoprofen, toward human and rat liver UGTs. Ketoprofen acylglucuronide irreversibly inhibited the glucuronidation of 1-naphthol and 2-naphthol catalyzed by human liver microsomes or by the recombinant rat liver isoform, UGT2B1, which is the main isoform involved in the glucuronidation of the drug. A decrease of about 35% in the glucuronidation of 2-naphthol was observed when ketoprofen acylglucuronide was produced in situ in cultured V79 cells expressing UGT2B1. Inhibition was always associated with the formation of microsomal protein-ketoprofen adducts. The presence of these covalent adducts within the endoplasmic reticulum of cells expressing UGT2B1 was demonstrated following addition of ketoprofen to culture medium by immunofluorescence microscopy with antiketoprofen antibodies. Immunoblots of liver microsomes incubated with ketoprofen acylglucuronide and probed with antiketoprofen antibodies revealed the presence of several protein adducts; among those was a major immunoreactive protein at 56 kDa, in the range of the apparent molecular mass of UGTs. The adduct formation partially prevented the photoincorporation of the UDP-glucuronic acid (UDP-GlcUA) analog, [beta-32P]5N3UDP-GlcUA, on the UGTs, suggesting that ketoprofen glucuronide covalently reacted with the UDP-GlcUA binding domain. Finally, UGT purification from rat liver microsomes incubated with ketoprofen glucuronide led to the isolation of UGT adducts recognized by both anti-UGT and antiketoprofen antibodies, providing strong evidence that UGTs are targets of this metabolite.     

Protein-loaded poly(epsilon-caprolactone) microparticles. I. Optimization of the preparation by (water-in-oil)-in water emulsion solvent evaporation.

The aim of this work was to optimize protein entrapment in pure poly(epsilon-caprolactone) (PCL) microparticles (MP) using the (water-in-oil)-in water solvent evaporation technique and bovine serum albumin (BSA) as drug model. Therefore, the preparative variables such as polymer solvent, protein/polymer ratio, polymer molecular weight, internal aqueous/organic phases ratio, organic/external aqueous phase ratio, and nature of the emulsifier were evaluated on microparticle characteristics such as BSA entrapment, entrapment efficiency, size and morphology. The in vitro release profiles of BSA from such MP in two different media with or without sodium dodecyl sulphate (SDS) were investigated. In optimum conditions, smooth and spherical pure PCL MP with high encapsulation efficiency (50.29 +/- 5.01%) were prepared. The release profiles of BSA in the release media were significantly different and faster in the presence of SDS. Moreover, they exhibited a relatively low burst effect after 24h (<30%) followed by a continuous release over 28 days. Due to PCL's numerous desirable characteristics, such MP could be an exciting alternative for the controlled release of proteinaceous compounds.     

Existence of different subtypes of nicotinic acetylcholine receptors in the rat habenulo-interpeduncular system.

Neuronal nicotinic ACh receptors (nAChRs) are present in the rat medial habenula (MHB) and interpeduncular nucleus (IPN), two brain regions connected through the fasciculus retroflexus (FR). The goal of the present study was to compare the electrophysiological and pharmacological characteristics of nAChRs located at pre- and postsynaptic sites within the MHB-IPN system. nAChRs located on the soma of IPN neurons were studied using patch-clamp techniques and a preparation of acutely isolated neurons. Whole-cell currents evoked by Ach and nicotine showed an intense rectification at positive membrane potentials. nAChR channels were relatively nonselective for cations, had a unitary conductance of 35 pS, and were activated by several nicotinic agonists with the following rank order: cytisine greater than ACh greater than nicotine greater than dimethylphenylpiperazinium (DMPP). They were blocked by mecamylamine, hexamethonium, curare, and dihydro-beta-erythroidine (DHBE), but were insensitive to alpha-bungarotoxin and neuronal bungarotoxin. In contrast, nAChRs recorded on the soma of MHB neurons under equivalent experimental conditions exhibited different characteristics for single-channel conductance and agonist and antagonist sensitivity. The pharmacological properties of presynaptic nAChRs in the IPN were analyzed in a rat brain slice preparation. Stimulation of the FR produced a presynaptic afferent volley recorded in the rostral subnucleus of the IPN. Nicotinic agonists decreased the amplitude of the afferent volley with different efficacies: nicotine greater than cytisine greater than ACh greater than DMPP. The action of nicotine was insensitive to alpha-bungarotoxin and to neuronal bungarotoxin, but was blocked by mecamylamine, hexamethonium, curare, and DHBE, with IC50 values different from those reported for IPN postsynaptic nAChRs. This study thus demonstrates the functional diversity of nAChRs in the rat CNS.