Effects of inhibiting mTOR with rapamycin on behavior, development, neuromuscular physiology and cardiac function in larval Drosophila.

Rapamycin and other mTOR inhibitors are being heralded as possible treatments for many human ailments. It is currently being utilized clinically as an immunomodulator after transplantation procedures and as a treatment for certain forms of cancer, but it has numerous potential clinical indications. Some studies have shown profound effects on life cycle and muscle physiology, but these issues have not been addressed in an organism undergoing developmental processes. This paper fills this void by examining the effect of mTOR inhibition by rapamycin on several different qualities of larval Drosophila Various dosages of the compound were fed to second instar larvae. These larvae were monitored for pupae formation to elucidate possible life cycle effects, and a delay to pupation was quantified. Behavioral deficits were documented in rapamycin-treated larvae. Electrophysiological measurements were taken to discern changes in muscle physiology and synaptic signaling (i.e. resting membrane potential, amplitude of excitatory post-synaptic potentials, synaptic facilitation). Pupation delay and effects on behavior that are likely due to synaptic alterations within the central nervous system were discovered in rapamycin-fed larvae. These results allow for several conclusions as to how mTOR inhibition by rapamycin affects a developing organism. This could eventually allow for a more informed decision when using rapamycin and other mTOR inhibitors to treat human diseases, especially in children and adolescents, to account for known side effects.

Targeted Delivery of TNF Potentiates the Antibody-Dependent Cell-Mediated Cytotoxicity of an Anti-Melanoma Immunoglobulin.

The recombinant murine IgG2a antibody TA99, directed against a melanoma antigen, was used to study combination modalities that potentiate antibody-dependent cell cytotoxicity. As previously reported, IgG2a(TA99) was extremely efficacious in preventing the growth of B16 lung metastases. However, the same antibody mediated only minimal tumor growth retardation when used to treat established neoplastic masses. The therapeutic activity of IgG2a(TA99) could be substantially enhanced by co-administration with an antibody-cytokine fusion (TA99-murine tumor necrosis factor [mTNF]), consisting of the TA99 antibody in single-chain variable fragment format fused to murine TNF. This fusion protein efficiently killed endothelial cells in vitro and displayed only minimal activity against B16 melanoma cells. In vivo, TA99-mTNF boosted the influx of natural killer cells and macrophages into B16 melanoma lesions. Therapy studies with two different administration schedules showed that the combination of TA99-mTNF and IgG2a(TA99) was superior to the individual products used as single agents. The combination treatment converted most of the tumor mass into a necrotic lesion, but a vital tumor rim eventually regrew, even when dacarbazine was included in the therapeutic regimen. The treatment modality described in this article may be applicable to the treatment of melanoma patients, given the specificity of the gp75 antigen and its conservation across species.

New insights into the acute actions from a high dosage of fluoxetine on neuronal and cardiac function: Drosophila, crayfish and rodent models.

The commonly used mood altering drug fluoxetine (Prozac) in humans has a low occurrence in reports of harmful effects from overdose; however, individuals with altered metabolism of the drug and accidental overdose have led to critical conditions and even death. We addressed direct actions of high concentrations on synaptic transmission at neuromuscular junctions (NMJs), neural properties, and cardiac function unrelated to fluoxetine's action as a selective 5-HT reuptake inhibitor. There appears to be action in blocking action potentials in crayfish axons, enhanced occurrences of spontaneous synaptic vesicle fusion events in the presynaptic terminals at NMJs of both Drosophila and crayfish. In rodent neurons, cytoplasmic Ca(2+) rises by fluoxetine and is thapsigargin dependent. The Drosophila larval heart showed a dose dependent effect in cardiac arrest. Acute paralytic behavior in crayfish occurred at a systemic concentration of 2mM. A high percentage of death as well as slowed development occurred in Drosophila larvae consuming food containing 100µM fluoxetine. The release of Ca(2+) from the endoplasmic reticulum in neurons and the cardiac tissue as well as blockage of voltage-gated Na(+) channels in neurons could explain the effects on the whole animal as well as the isolated tissues. The use of various animal models in demonstrating the potential mechanisms for the toxic effects with high doses of fluoxetine maybe beneficial for acute treatments in humans. Future studies in determining how fluoxetine is internalized in cells and if there are subtle effects of these mentioned mechanisms presented with chronic therapeutic doses are of general interest.