Ceramide analogues in apoptosis: a new strategy for anticancer drug development

A survey on the role played by ceramide within the sphingolmyelin pathway is here reported, taking into account its importance as an intracellular effector molecule in apoptosis. Recently, several analogs of ceramide, able to pass the cell membrane and then to induce apoptosis, have been developed as a new potential approach in anticancer therapy.     

A Reappraisal of Drug Release Laws Using Monte Carlo Simulations: The Prevalence of the Weibull Function

Purpose. To verify the Higuchi law and study the drug release from cylindrical and spherical matrices by means of Monte Carlo computer simulation. Methods. A one-dimensional matrix, based on the theoretical assumptions of the derivation of the Higuchi law, was simulated and its time evolution was monitored. Cylindrical and spherical three-dimensional lattices were simulated with sites at the boundary of the lattice having been denoted as leak sites. Particles were allowed to move inside it using the random walk model. Excluded volume interactions between the particles was assumed. We have monitored the system time evolution for different lattice sizes and different initial particle concentrations. Results. The Higuchi law was verified using the Monte Carlo technique in a one-dimensional lattice. It was found that Fickian drug release from cylindrical matrices can be approximated nicely with the Weibull function. A simple linear relation between the Weibull function parameters and the specific surface of the system was found. Conclusions. Drug release from a matrix, as a result of a diffusion process assuming excluded volume interactions between the drug molecules, can be described using a Weibull function. This model, although approximate and semiempirical, has the benefit of providing a simple physical connection between the model parameters and the system geometry, which was something missing from other semiempirical models.     

Computer Simulation of Convective and Diffusive Transport of Controlled-Release Drugs in the Vitreous Humor

Purpose. Biodistribution of drugs in the eye is central to the efficacy of pharmaceutical ocular therapies. Of particular interest to us is the effect of intravitreal transport on distribution of controlled-released drugs within the vitreous. Methods. A computer model was developed to describe the three-dimensional convective-diffusive transport of drug released from an intravitreal controlled release source. Unlike previous studies, this work includes flow of aqueous from the anterior to the posterior of the vitreous. The release profile was based on in vitro release of gentamicin from poly(L-lactic acid) microspheres into vitreous. Results. For small drugs, convection plays a small role, but for large (slower diffusing) drugs, convection becomes more important. For the cases studied, the predicted ratio of drug reaching the retina to drug cleared by the aqueous humor was 2.4 for a small molecule but 13 for a large molecule. Transport in neonatal mouse eye, in contrast, was dominated by diffusion, and the ratio decreased to 0.39. Conclusions. The interaction among convection, diffusion, and geometry causes significant differences in biodistribution between large and small molecules or across species. These differences should be considered in the design of delivery strategies or animal studies.     

Pairing elevation of [cyclic GMP] with inhibition of PKA produces long-term depression of glutamate release from isolated rat hippocampal presynaptic terminals

Data suggest both presynaptic and postsynaptic changes contribute to activity-dependent long-term synaptic plasticity. We have shown that pairing elevation of intracellular [cyclic GMP], using the type V phosphodiesterase inhibitor zaprinast, with inhibition of cyclic AMP-dependent protein kinase (PKA), is sufficient to elicit chemical long-term depression (CLTD) of synaptic transmission at Schaffer collateral-CA1 and mossy fibre-CA3 synapses in rat hippocampus. CLTD does not require synaptic activity, and selective postsynaptic drug injections do not affect it, suggesting it is presynaptically induced and expressed. To directly evaluate this hypothesis, we tested whether CLTD of transmitter release can be expressed in isolated presynaptic nerve terminals. Presynaptic nerve terminals (synaptosomes) were isolated from rat hippocampi by Percoll density gradient centrifugation. Synaptosomes were loaded with [3H]glutamate, and basal and depolarisation-induced release of [3H]glutamate measured in control medium versus medium containing zaprinast (20 microm) plus or minus the PKA inhibitor H-89 (10 microm). Zaprinast produced a significant decrease in basal [3H]glutamate release. However, only combining zaprinast with H-89 significantly depressed K+-evoked [3H]glutamate release. After a 20-min drug washout, basal release returned to normal in all conditions, but K+-evoked [3H]glutamate release was persistently reduced only by the combination of zaprinast plus H-89. Long-term reduction of [3H]glutamate release from synaptosomes was completely prevented by the PKG inhibitor KT5823 (5 microm). These data demonstrate the existence of a presynaptic, cyclic GMP-PKG dependent cascade capable of expressing LTD of glutamate release from isolated hippocampal nerve terminals.     

Conditional involvement of striatal serotonin3 receptors in the control of in vivo dopamine outflow in the rat striatum

Serotonin3 (5-HT3) receptors can affect motor control through an interaction with the nigrostriatal dopamine (DA) neurons, but the neurochemical basis for this interaction remains controversial. In this study, using in vivo microdialysis, we assessed the hypothesis that 5-HT3 receptor-dependent control of striatal DA release is conditioned by the degree of DA and/or 5-HT neuron activity and the means of DA release (impulse-dependent vs. impulse-independent). The different DA-releasing effects of morphine (1 and 10 mg/kg), haloperidol (0.01 mg/kg), amphetamine (1 and 2.5 mg/kg), and cocaine (10 and 20 mg/kg) were studied in the striatum of freely moving rats administered selective 5-HT3 antagonists ondansetron (0.1 mg/kg) or MDL 72222 (0.03 mg/kg). Neither of the 5-HT3 antagonists modified basal DA release by itself. Pretreatment with ondansetron or MDL 72222 reduced the increase in striatal DA release induced by 10 mg/kg morphine but not by 1 mg/kg morphine, haloperidol, amphetamine or cocaine. The effect of 10 mg/kg morphine was also prevented by intrastriatal ondansetron (1 microm) administration. Reverse dialysis with ondansetron also reduced the increase in DA release induced by the combination of haloperidol and the 5-HT reuptake inhibitor citalopram (1 mg/kg). Considering the different DA and 5-HT-releasing properties of the drugs used, our results demonstrate that striatal 5-HT3 receptors control selectively the depolarization-dependent exocytosis of DA only when central DA and 5-HT tones are increased concomitantly.     

Blockade of noradrenergic receptors in the basolateral amygdala impairs taste memory

In conditioned taste aversion (CTA), a subject learns to associate a novel taste (conditioned stimulus, CS) with visceral malaise (unconditioned stimulus, US). Considerable evidence indicates that the noradrenergic system in the amygdala plays an important role in memory consolidation for emotionally arousing experiences. The specific aim of the present set of experiments was to determine the involvement of noradrenergic activity in the basolateral amygdala (BLA) during the US presentation and consolidation of CTA as well as during the consolidation of a nonaversive/incidental gustatory memory. Selective bilateral microinfusions of the beta-adrenergic antagonist propranolol administered into the BLA immediately before intraperitoneal (i.p.) lithium chloride (LiCl) injections disrupted CTA memory. Additionally, propranolol infused into the BLA immediately after a pre-exposure to the saccharin (CS) significantly attenuated latent inhibition. The present findings indicating that alterations in noradrenergic function in the BLA affect taste memory formation, provide additional evidence that the BLA plays a critical role in modulating the consolidation of memory and that the influence is mediated by interactions with other brain regions that support memory for different kinds of experiences.     

Fluoxetine depresses glutamate exocytosis in the rat cerebrocortical nerve terminals (synaptosomes) via inhibition of P/Q-type Ca2+ channels

Fluoxetine, an antidepressant that is used clinically in the treatment of mood disorders, is a selective serotonin reuptake inhibitor. In the present study we investigated the effects of fluoxetine on 4-aminopyridine (4AP)-evoked glutamate release in cerebrocortical nerve terminals (synaptosomes). Fluoxetine suppressed the release of glutamate evoked by 4AP in a concentration-dependent manner. This effect was associated with a reduction in the depolarization-evoked increase in cytosolic free calcium levels in the absence of significant effect on the synaptosomal membrane potential. In addition, both ionomycin- and sucrose-evoked glutamate releases were not affected by fluoxetine, indicating that fluoxetine-mediated inhibition of glutamate release is not a direct effect on the exocytotic machinery. Furthermore, the inhibitory action of fluoxetine was completely abolished in synaptosomes pretreated with P/Q type Ca(2+) channel blocker omega-agatoxin IVA (omega-AgTX IVA) or protein kinase C (PKC) stimulator 4beta-phorbol 12, 13-dibutyrate (PDBu). These results suggest that, in cerebrocortical nerve terminals, fluoxetine inhibits glutamate release through the suppression of P/Q type Ca(2+) channel activity. The presynaptic action of fluoxetine is mediated by a PKC-sensitive signaling pathway. Synapse 48:170-177, 2003.     

Presynaptic quantal plasticity: Katz's original hypothesis revisited

Changes in the amplitudes of signals conveyed at synaptic contacts between neurons underlie many brain functions and pathologies. Here we review the possible determinants of the amplitude and plasticity of the elementary postsynaptic signal, the miniature. In the absence of a definite understanding of the molecular mechanism releasing transmitters, we investigated a possible alternative interpretation. Classically, both the quantal theory and the vesicle theory predict that the amount of transmitter producing a miniature is determined presynaptically prior to release and that rapid changes in miniature amplitude reflect essentially postsynaptic alterations. However, recent data indicates that short-term and long-lasting changes in miniature amplitude are in large part due to changes in the amount of transmitter in individual released packets that show no evidence of preformation. Current representations of transmitter release derive from basic properties of neuromuscular transmission and endocrine secretion. Reexamination of overlooked properties of these two systems indicate that the amplitude of miniatures may depend as much, if not more, on the Ca(2+) signals in the presynaptic terminal than on the number of postsynaptic receptors available or on vesicle's contents. Rapid recycling of transmitter and its possible adsorption at plasma and vesicle lumenal membrane surfaces suggest that exocytosis may reflect membrane traffic rather than actual transmitter release. This led us to reconsider the disregarded hypothesis introduced by Fatt and Katz (1952; J Physiol 117:109-128) that the excitability of the release site may account for the "quantal effect" in fast synaptic transmission. In this case, changes in excitability of release sites would contribute to the presynaptic quantal plasticity that is often recorded.     

Bovine mastitis and intramammary drug delivery: review and perspectives

Intramammary infections (IMIs) represent a major feature in bovine pathology. The treatment of IMIs concern antimicrobial substances. Therapeutic strategies involve administration of immediate release formulations during lactation with or without long-acting formulations during the dry period. Current treatments are not very successful and cure rates are poor, especially towards Staphylococcus aureus which is responsible for chronic infections and huge economic losses. New strategies have recently been investigated. These include particular immunomodulators like lysostaphin or cytokines, and novel formulations (e.g. liposomes, microparticles or nanoparticles) that allow uptake of the active component by phagocytes and thus prolong an enhanced antibacterial activity.