Apolipoprotein E*3-Leiden transgenic mice as a test model for hypolipidaemic drugs

PURPOSE: We report an investigation into the distribution of proteoglycans (PGs) in normal, organ-cultured and dextran-treated human corneas. METHODS: Immunogold labeling was carried out at the electron microscope level to localize keratan sulphate (KS), chondroitin sulphate (CS), and heparan sulphate (HS) PGs. RESULTS: High levels of labeling for CS was found in the epithelium, endothelium, and keratocytes, with light labelling present in the basement membranes and the corneal stroma. Labeling for HS was present in the epithelium, endothelium, and keratocytes, with intense labeling present at the endothelium/Descemet's membrane interface and the epithelium/Bowman's layer interface. Large filaments were also observed in these regions in cuprolinic blue-stained specimens. Keratan sulphate was present at high levels in the stroma and the basement membranes with low levels present within the keratocytes, epithelium, and endothelium. The pattern of KS labeling along the collagen fibrils in the stroma sometimes showed evidence of periodicity. Organ-cultured corneas had extensive collagen-free "lakes," the interior of which immunolabeled positively for KS and showed staining with cuprolinic blue. The lakes were greatly reduced in the dextran-treated samples. CONCLUSION: This investigation determined the ultrastructural distribution of KS, CS, and HS PGs in human cornea and showed that organ culture is associated with a change in distribution of stromal PGs.     

The effects of age and gender on the single dose pharmacokinetics of avitriptan administered to healthy volunteers

The effects of age and gender on the single dose pharmacokinetics of avitriptan and its three metabolites were assessed in 15 young men, 15 young women, 15 elderly men and 15 elderly women. Avitriptan was administered as a 150-mg capsule after a 10-hour fast and serial plasma and urine samples were collected up to 36 hours after the dose. Plasma samples were analyzed for avitriptan and its metabolites, N-desmethyl avitriptan (ND048), O-desmethyl avitriptan (OD048), and methoxypyrimidinyl piperazine (MPP). Urine samples were analyzed for only avitriptan and MPP. Avitriptan was well tolerated in all four groups. The drug was rapidly absorbed with a median time to maximum plasma concentration (tmax) between 0.5 and 1.5 hours. No significant gender-related differences were found in the maximum plasma concentration (Cmax) and area under the concentration-time curve extrapolated to infinity (AUC0-infinity) of avitriptan. Renal clearance of avitriptan was significantly smaller in young women compared with young men, but this is clinically not relevant because only 2% to 3% of the total dose is excreted unchanged. Compared with the young volunteers, mean Cmax was approximately 50% higher in the elderly but there was no difference in the AUC0-infinity between the 2 age groups. Plasma concentrations of ND048, OD048, and MPP were each 50 to 100 fold lower than those of avitriptan. Hence some age- and gender-related differences found in the pharmacokinetics of avitriptan metabolites are probably not relevant in the assessment of overall safety and efficacy of avitriptan. Based on the pharmacokinetics and tolerability, no age or gender-related dose adjustment is necessary for avitriptan.     

Addisonian crisis provoked by levothyroxine substitution therapy

A women aged 36 with a positive family anamnesis for autoimmune endocrine diseases and a history of thyroid diseases, developed major complaints of general malaise, orthostatic hypotension and loss of appetite after the start of a treatment with levothyroxin because of (sub)clinical hypothyroidism. She was found to suffer from primary adrenocortical insufficiency masked by excessive use of liquorice and a lowered metabolism, but which via the suppletion with thyroid hormone had led to an addisonian crisis.     

Modification of hypoxia-induced injury in cultured rat astrocytes by high levels of glucose

BACKGROUND AND PURPOSE: Preexisting hyperglycemia exacerbates central nervous system injury after transient global and focal cerebral ischemia. Increased anaerobic metabolism with resultant lactic acidosis has been shown to cause the hyperglycemic, neuronal injury. The contribution of astrocytes in producing lactic acidosis under hyperglycemic/ischemic conditions is unclear, whereas the protective role of astrocytes in ischemic-induced neuronal injury has been documented. The ability of astrocytes to maintain energy status and ion homeostasis under hyperglycemic conditions could ultimately reduce neuronal injury. Therefore, we determined the effects of increased glucose concentrations on glucose utilization, lactate production, extracellular pH, and adenosine triphosphate concentrations in hypoxia-treated astrocyte cultures. METHODS: Primary astrocytes were prepared from neonatal rat cerebral cortices. After 35 days in vitro, cultures were incubated with 0-60 mmol/L glucose and subjected to hypoxic conditions at 95% N2/5% CO2 for 24 hours. In addition, under high-glucose conditions (30 mmol/L), astrocytes were exposed to up to 72 hours of hypoxia. Determination of lactate dehydrogenase efflux, adenosine triphosphate concentrations, and extracellular lactate concentrations defined astrocyte status. Equiosmolar levels of mannitol were added in place of high glucose concentrations to distinguish hyperosmotic effect. RESULTS: When physiological concentrations of glucose (7.5 mmol/L) or lower concentrations were used, significant cell damage occurred with 24 hours of hypoxia, as determined by increased efflux of lactate dehydrogenase and loss of cell protein. When higher glucose concentrations (15-60 mmol/L) were used, efflux of lactate dehydrogenase was similar to that observed in normoxic cultures, despite an increased utilization of glucose. Lactate concentrations in the media at low or normal glucose concentrations exceeded normoxic levels, but higher glucose concentrations (15-30 mmol/L) failed to increase lactate levels further. Values of adenosine triphosphate for hypoxic astrocytes treated with high glucose concentrations were significantly higher than those of astrocytes with zero or low glucose levels. In cultures exposed to hypoxia and high glucose levels (30 mmol/L), no cellular injury was observed before 48 hours of hypoxia. Lactate concentrations in the media increased during the first 24 hours of hypoxia and reached steady state. The pH of the media decreased to 6.4 after 24 hours and 5.5 at 48 hours. The latter pH was concomitant with a marked increase in extracellular lactate dehydrogenase activity. Hyperosmotic mannitol failed to protect cultured astrocytes against hypoxia. CONCLUSIONS: Hypoxic injury to mature astrocytes was reduced by the presence of 15-60 mmol/L glucose in the medium during 24-30 hours of hypoxia. Injury occurred when the pH of the medium was < 5.5. This protection was not afforded by the hyperosmotic effect of high glucose concentrations, nor was the hypoxic injury at later time periods with 30 mmol/L glucose mediated solely by lactate accumulation.     

The Glu318Gly mutation of the presenilin-1 gene does not necessarily cause Alzheimer's disease

In early-onset familial Alzheimer's disease (AD) pathogenic mutations have been found in the amyloid precursor protein (APP) gene and in the presenilin (PS)-1 and PS-2 genes. We screened for mutations in these genes in 20 patients with familial AD from the Finnish population. In addition, we sampled 41 sporadic AD patients and 59 controls to test for mutations identified in our familial AD cases. We detected an A-to-G transition in the PS-1 gene, resulting in a glutamic acid (Glu)-to-glycine (Gly) substitution at codon 318 in 2 familial and 2 sporadic AD patients. The Glu318Gly mutation has previously been reported to cause AD. We also found the Glu318Gly mutation in 4 healthy aged controls (range, 74-87 years). We thus conclude that the mutation is most likely a rare polymorphism not related to AD.     

Potentiation of the time-dependent, antidepressant-induced changes in the agonistic behaviour of resident rats by the 5-HT1A receptor antagonist, WAY-100635

Acute and chronic antidepressant drug treatments respectively decrease and increase the aggressive behaviour of resident rats during encounters with unfamiliar conspecifics. We have now examined the effect of the 5-hydroxytryptamine1A receptor antagonist, WAY-100635, on fluoxetine-, paroxetine- or venlafaxine-induced changes in aggression. WAY-100635 (0.1 mg/kg), which did not modify behaviour when given alone, potentiated the venlafaxine (5.54 mg/kg)-induced reduction in aggression after acute treatment and, during chronic treatment, accelerated the fluoxetine (0.34 mg/kg/day)-induced increase in aggression, from day 5 to day 2. A similar change in time course was seen with paroxetine (0.33 mg/kg/day), although the increase in aggression was smaller. Venlafaxine (5.54 mg/kg/day, alone or co-administered with WAY-100635) increased aggression by day 2. During chronic treatment, therefore, venlafaxine, at the dose used, had a more rapid onset of action than either fluoxetine or paroxetine, whereas the fluoxetine- and paroxetine-, but not the venlafaxine-, induced increase in aggression was accelerated by WAY-100635. These studies further support the hypothesis that selective blockade of the 5-hydroxytryptamine1A receptor augments the effects of antidepressant drugs in an animal model predictive of antidepressant activity, presumably by concomitant blockade of the somatodendritic 5-hydroxytryptamine1A autoreceptor-mediated negative feedback system of serotonergic neurones.     

Neuronal spike activity in the nucleus accumbens of behaving rats during ethanol self-administration

Many lines of evidence support the importance of the nucleus accumbens (NAC) for ethanol-reinforced behavior. The nature of the neuronal activity that occurs in this region during ethanol self-administration is not known. We recorded from ensembles of single-units primarily located within the shell of the NAC during operant responding for oral ethanol solutions by well-trained rats. Of 90 units recorded from seven sessions from seven rats, 41 (46%) did not exhibit significant changes in relation to the experimental events. Of the 49 units (54%) that did exhibit significant phasic changes, alterations in firing rate occurred in relation to the following experimental events: operant response (63%), tone stimulus (20%), and ethanol delivery (63%). In addition, changes in spike activity during the intervals between the three experimental events were noted in 33% of the units. Most units (55% of responsive units) responded to multiple experimental events. Thus different but overlapping populations of neurons in the NAC represent each event that occurs along the temporal dimension of a single trial performed to obtain ethanol reward. The data suggest that the NAC plays a crucial role in linking together conditioned and unconditioned internal and external stimuli with motor plans to allow for ethanol-seeking behavior to occur.     

Biosynthesis of scarab beetle pheromones

Chemical communication in scarab beetles (Coleoptera: Scarabaeidae) is achieved with a wide variety of pheromones, but one typical structure is the gamma-lactone having a long unsaturated hydrocarbon chain. Several species utilize (R, Z)-5-(-)-(oct-1-enyl)-oxacyclopentan-2-one (buibuilactone), (R, Z)-5-(-)-(dec-1-enyl)-oxacyclopentan-2-one and (S, Z)-5-(+)-(dec-1-enyl)-oxacyclopentan-2-one [(R)-japonilure and (S)-japonilure]. Using deuterated precursors, we have demonstrated that these compounds are biosynthesized from fatty acids. (9, 10-d4)-Palmitic acid, (9,10-d4)-stearic acid, (9,10-d2)-palmitoleic acid, (9,10-d2)-oleic acid, (9,10-d2)-8-hydroxypalmitoleic acid and (9,10-d2)-8-hydroxyoleic acid were readily incorporated by female Anomala cuprea into the pheromone molecules, while (Z)-(5, 6-d2)-5-dodecenoic acid and (Z)-(5,6-d2)-5-tetradecenoic acid were not. Therefore, the reaction pathway starts from saturated fatty acids, involves their desaturation, followed by 8-hydroxylation, chain shortening and cyclization. The products obtained from racemic (9,10-d2)-8-hydroxypalmitoleic acid and (9,10-d2)-8-hydroxyoleic acid were also racemic, implying that the steps following hydroxylation were not stereospecific. Perdeuterated palmitic acid was applied to disclose the mechanism of the unique hydroxylation reaction. Retention of all deuterium atoms implied that this reaction was a direct process mediated by a specific fatty acid hydroxylase, and preceding desaturation or epoxidation was not involved.