A coupled assay for histidine decarboxylase: in vivo turnover of this enzyme in mouse brain

Abstract

A sensitive coupled assay for histidine decarboxylase has been developed. This method involved conversion of [3H]histidine into [3H]histamine by the enzyme sample, with methylation of this product in situ, catalysed by the enzyme histamine N-methyltransferase, to yield [3H]N-tele-methylhistamine. The radioactive product was separated from the substrate by (i) extraction into chloroform, (ii) ion-exchange chromatography and (iii) liquid cation-exchange extraction. The "no tissue" assay blank comprised 0.0007% of the substrate radioactivity. Sample material with a histidine decarboxylase activity of as little as 0.14 fmol/min/ml (measured at 1 microM histidine) gave double the blank value. More than 50 assays could be performed in one day. This assay was used to determine the in vivo changes in mouse brain histidine decarboxylase activity following irreversible inhibition with (+) alpha-fluoromethylhistidine (alpha-FMH). From the time course of recovery of enzyme activity the half-life of histidine decarboxylase in vivo was calculated to be 53 h.

The Histamine H1 Receptor Participates in the Increased Dorsal Telencephalic Neurogenesis in Embryos from Diabetic Rats

“…A modified method was used to measure the HDC activity (Keeling et al, 1984 ; Barnes and Hough, 2002 ; Shoji et al, 2006 ). Five hundred milligrams of mesencephalon/rhombencephalon tissue (where fetal histaminergic transitory neurons reside) was obtained from E14 and E16, or from placenta at E14.…”

Quantitative Radioisotopic Determination of Histidine Decarboxylase Using High-Performance Liquid Chromatography

“…The blanks are critical in this assay, because the percentage of histidine converted to histamine is typically very low, and thus minor contaminants could greatly impair the reproducibility of results. Keeling et al (8) described a radioisotopic method of histidine decarboxylase determination in a paper that critically addressed the high level of dpm in the blanks of the original method developed by Baudry et al (7). However, Keeling's method was complex and also used ion-exchange gravity columns as a final step of purification.…”

“…Also, given the low rate of transformation of substrate into histamine (0.1-0.5% in rat brain homogenates), the sensitivity of this approach requires a strict separation between histamine and the excess histidine. In previously published procedures, the purification of both commercial radiolabeled histidine and enzymatically produced histamine has been accomplished using cation-exchange gravity columns (Amberlite CG-50) (7,8). Unfortunately, these columns do not provide effective purification of either substrate or product, giving rise to high background values and variable results.…”

The polyamine and histamine metabolic interplay in cancer and chronic inflammation

“…The key enzymes involved in both metabolic pathways, in particular ODC, SAM-decarboxylase (SAMDC), SSAT and HDC, are present in low abundance, exhibit rapid turnover and are under complex transcriptional and posttranscriptional regulation. As their half-lives are very short [9,10], their expression levels can change by several orders of magnitude very quickly in response to different types of stimuli. A family of proteinaceous inhibitors, called antizymes, regulates ODC degradation and thereby contribute to the control of polyamine homeostasis (reviewed in [11]).…”