A cholecystokinin (CCK)-inactivating peptidase was purified and identified as a membrane-bound isoform of tripeptidyl peptidase II (EC 3.4.14.10), a cytosolic subtilisin-like peptidase of previously unknown functions. The peptidase was found in neurons responding to cholecystokinin, as well as in non-neuronal cells. Butabindide, a potent and specific inhibitor, was designed and shown to protect endogenous cholecystokinin from inactivation and to display pro-satiating effects mediated by the CCKA receptor.
Brain diseases such as autism and Alzheimer's disease (each inflicting >1% of the world population) involve a large network of genes displaying subtle changes in their expression. Abnormalities in intraneuronal transport have been linked to genetic risk factors found in patients, suggesting the relevance of measuring this key biological process. However, current techniques are not sensitive enough to detect minor abnormalities. Here we report a sensitive method to measure the changes in intraneuronal transport induced by brain-disease-related genetic risk factors using fluorescent nanodiamonds (FNDs). We show that the high brightness, photostability and absence of cytotoxicity allow FNDs to be tracked inside the branches of dissociated neurons with a spatial resolution of 12 nm and a temporal resolution of 50 ms. As proof of principle, we applied the FND tracking assay on two transgenic mouse lines that mimic the slight changes in protein concentration (∼30%) found in the brains of patients. In both cases, we show that the FND assay is sufficiently sensitive to detect these changes.
—Properties of the histamine‐forming enzyme in rat brain were studied, utilizing a sensitive fluorometric assay. The optimum pH was related to substrate concentration and found to be6·4 at 10−2m‐histidine; the apparent Km was about 4·10−4m; enzyme activity was inhibited by α‐hydrazino ‐histidine and brocresine but was not affected by α‐methyl DOPA or benzene. These different data suggest that the 'specific’histidine decarboxylase (EC 4.1.1.22)—and not the aromatic l‐aminoacid decarboxylase—is involved. Determination of enzyme activity and histamine level in different areas of the rat brain revealed important regional differences, the two values being roughly parallel.
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