Pharmacological characteristics of protein synthesis inhibitors by radioactive leucine incorporation in rat hippocampal slices: experimental evidence and pre-clinical implications

• Protein synthesis inhibitors (PSIs) constitute a major tool to validate the hypothesis of protein synthesis-dependent phase of synaptic plasticity and memory consolidation. However, several reports have showed inconsistent findings about the effect of these drugs on behavioral learning and synaptic plasticity. Testing the potencies of these drugs is hence crucial for validating such negative findings and in planning future studies. It is also necessary to examine the dose dependence, onset dynamics and reversibility, and possible effects on basal proteins. Here we used the labeled leucine as marker for the newly synthesized proteins. The fraction of leucine incorporation, following 50 min of pre-incubation, was compared between two groups of slices: a PSI-treated and a control group. Both anisomycin and cycloheximide revealed a dose-dependent but time-independent manner of inhibition reaching over 92% at concentrations well below those used in previous experiments which revealed effects on synaptic plasticity and learning. Surprisingly, washout of a “reversible” inhibitor, anisomycin was not followed by rapid reversibility of the action of the drug, the case that differs with cycloheximide. Interestingly, emetine revealed a time-dependent inhibition of protein synthesis, where levels above 80% needed drug pre-incubation for as long as 90 min. Since the duration of labeling relates to the half-life of the proteins, short-time labeling as used in this study will result in radioactivity incorporation into short-lived proteins and proteins that are synthesized in large quantities. We therefore studied the availability of newly synthesized proteins at 8-10 h following leucine incorporation. The results revealed virtually the same protein content as in slices retrieved for analysis immediately following the labeling period, indicating that the main pool of the newly synthesized proteins is of intracellular long-lived pool. This likely reflects a stable metabolic state of our prepared slices. These findings challenge current idea on the role of de novo protein synthesis in synaptic plasticity as well as brain changes underlying several neurological and psychiatric disorders.

Subject headings

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinska och farmaceutiska grundvetenskaper -- Cell- och molekylärbiologi (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Basic Medicine -- Cell and Molecular Biology (hsv//eng)

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinska och farmaceutiska grundvetenskaper -- Neurovetenskaper (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Basic Medicine -- Neurosciences (hsv//eng)

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinska och farmaceutiska grundvetenskaper -- Farmakologi och toxikologi (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Basic Medicine -- Pharmacology and Toxicology (hsv//eng)

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