Amitriptyline and imipramine inhibit the release of acetylcholine from parasympathetic nerve terminals in the rat iris

The electrically stimulated release of [3H]acetylcholine from the parasympathetic nerve terminals of the rat iris in vitro is increased in a dose-dependent manner by scopolamine but is decreased by the tricyclic antidepressants amitriptyline and imipramine. The increased release in the presence of scopolamine seems to be due to the blockade of a presynaptic muscarinic autoreceptor that, in the drug-free state, inhibits the release of acetylcholine. However, at drug concentrations that should have comparable antimuscarinic potency, the antidepressants inhibit the release of acetylcholine. This suggests that the anticholinergic side effects of the antidepressants may be due to the reduced release of acetylcholine from parasympathetic nerve terminals as well as a possible direct postsynaptic muscarinic receptor blocking action. Whatever the mechanism of this action, the antidepressants do not have the same effect as scopolamine at the presynaptic muscarinic autoreceptor in the rat iris.     

Cathepsin L participates in the production of neuropeptide Y in secretory vesicles, demonstrated by protease gene knockout and expression

Neuropeptide Y (NPY) functions as a peptide neurotransmitter and as a neuroendocrine hormone. The active NPY peptide is generated in secretory vesicles by proteolytic processing of proNPY. Novel findings from this study show that cathepsin L participates as a key proteolytic enzyme for NPY production in secretory vesicles. Notably, NPY levels in cathepsin L knockout (KO) mice were substantially reduced in brain and adrenal medulla by 80% and 90%, respectively. Participation of cathepsin L in producing NPY predicts their colocalization in secretory vesicles, a primary site of NPY production. Indeed, cathepsin L was colocalized with NPY in brain cortical neurons and in chromaffin cells of adrenal medulla, demonstrated by immunofluorescence confocal microscopy. Immunoelectron microscopy confirmed the localization of cathepsin L with NPY in regulated secretory vesicles of chromaffin cells. Functional studies showed that coexpression of proNPY with cathepsin L in neuroendocrine PC12 cells resulted in increased production of NPY. Furthermore , in vitro processing indicated cathepsin L processing of proNPY at paired basic residues. These findings demonstrate a role for cathepsin L in the production of NPY from its proNPY precursor. These studies illustrate the novel biological role of cathepsin L in the production of NPY, a peptide neurotransmitter, and neuroendocrine hormone.     

Human cathepsin V protease participates in production of enkephalin and NPY neuropeptide neurotransmitters

Proteases are required for processing precursors into active neuropeptides that function as neurotransmitters for cell-cell communication. This study demonstrates the novel function of human cathepsin V protease for producing the neuropeptides enkephalin and neuropeptide Y (NPY). Cathepsin V is a human-specific cysteine protease gene. Findings here show that expression of cathepsin V in neuroendocrine PC12 cells and human neuronal SK-N-MC cells results in production of (Met)enkephalin from proenkephalin. Gene silencing of cathepsin V by siRNA in human SK-N-MC cells results in reduction of (Met)enkephalin by more than 80%, illustrating the prominent role of cathepsin V for neuropeptide production. In vitro processing of proenkephalin by cathepsin V occurs at dibasic residue sites to generate enkephalin-containing peptides and an ～24-kDa intermediate present in human brain. Cathepsin V is present in human brain cortex and hippocampus where enkephalin and NPY are produced and is present in purified human neuropeptide secretory vesicles. Colocalization of cathepsin V with enkephalin and NPY in secretory vesicles of human neuroblastoma cells was illustrated by confocal microscopy. Furthermore, expression of cathepsin V with proNPY results in NPY production. These findings indicate the unique function of human cathepsin V for producing enkephalin and NPY neuropeptides required for neurotransmission in health and neurological diseases.