INMT - Indolethylamine N-methyltransferase

Before techniques of molecular biology were used to localize indolethylamine N-methyltransferase (INMT), characterization and localization went on a par: samples of the biological material where INMT is hypothesized to be active are subject to enzyme assay. 

Those enzyme assays are performed either with a radiolabeled methyl donor like (14C-CH3)SAM to which known amounts of unlabeled substrates like tryptamine are added or with addition of a radiolabeled substrate like (14C)NMT to demonstrate in vivo formation.

As qualitative determination of the radioactively tagged product of the enzymatic reaction is sufficient to characterize INMT existence and activity (or lack of), analytical methods used in INMT assays are not required to be as sensitive as those needed to directly detect and quantify the minute amounts of endogenously formed DMT (see DMT subsection below). The essentially qualitative method thin layer chromatography (TLC) was thus used in a vast majority of studies.

Also, robust evidence that INMT can catalyze transmethylation of tryptamine into NMT and DMT could be provided with reverse isotope dilution analysis coupled to mass spectrometry for rabbit and human lung during the early 1970s.

Selectivity rather than sensitivity proved to be an Achilles' heel for some TLC methods with the discovery in 1974–1975 that incubating rat blood cells or brain tissue with (14C-CH3)SAM and NMT as substrate mostly yields tetrahydro-β-carboline derivatives, and negligible amounts of DMT in brain tissue.

It is indeed simultaneously realized that the TLC methods used thus far in almost all published studies on INMT and DMT biosynthesis are incapable to resolve DMT from those tetrahydro-β-carbolines. 

These findings are a blow for all previous claims of evidence of INMT activity and DMT biosynthesis in avian and mammalian brain, including in vivo, as they all relied upon use of the problematic TLC methods: their validity is doubted in replication studies that make use of improved TLC methods, and fail to evidence DMT-producing INMT activity in rat and human brain tissues. 

Published in 1978, the last study attempting to evidence in vivo INMT activity and DMT production in brain (rat) with TLC methods finds biotransformation of radiolabeled tryptamine into DMT to be real but "insignificant".

Capability of the method used in this latter study to resolve DMT from tetrahydro-β-carbolines is questioned later.

To localize INMT, a qualitative leap is accomplished with use of modern techniques of molecular biology, and of immunohistochemistry. In humans, a gene encoding INMT is determined to be located on chromosome 7.

Northern blot analyses reveal INMT messenger RNA (mRNA) to be highly expressed in rabbit lung, and in human thyroid, adrenal gland, and lung.

Intermediate levels of expression are found in human heart, skeletal muscle, trachea, stomach, small intestine, pancreas, testis, prostate, placenta, lymph node, and spinal cord.

Low to very low levels of expression are noted in rabbit brain, and human thymus, liver, spleen, kidney, colon, ovary, and bone marrow.

INMT mRNA expression is absent in human peripheral blood leukocytes, whole brain, and in tissue from 7 specific brain regions (thalamus, subthalamic nucleus, caudate nucleus, hippocampus, amygdala, substantia nigra, and corpus callosum).

Immunohistochemistry showed INMT to be present in large amounts in glandular epithelial cells of small and large intestines.

In 2011, immunohistochemistry revealed the presence of INMT in primate nervous tissue including retina, spinal cord motor neurons, and pineal gland.

A 2020 study using in-situ hybridization, a far more accurate tool than the northern blot analysis, found mRNA coding for INMT expressed in the human cerebral cortex, choroid plexus, and pineal gland.