Analysis of proenkephalin A, proopiomelanocortin and protachykinin neuropeptides in human lumbar cerebrospinal fluid by reversed-phase high-performance liquid chromatography, radioimmunoassay and enzymolysis.
A comprehensive high-performance liquid chromatographic, radioimmunoassay, and enzymatic degradation scheme has been developed to analyze several intact neuropeptides and the corresponding peptides created by in vivo enzymolysis of precursors to study neuropeptides in human lumbar cerebrospinal fluid (CSF) and to test the hypothesis that defects in the metabolism (synthesis, degradation) of neuropeptide precursors, neuropeptides, and metabolites play a role in low back pain. CSF samples were obtained from three different patient groups: controls (C), whose low back pain was relieved without lidocaine; pharmacological responders (PR), whose pain was relieved by lidocaine and who were candidates for surgery; and pharmacological non-responders (PNR), whose pain was not relieved by lidocaine and a mid-thoracic anesthetic, and who were not candidates for surgery. The metabolic activity involved during synthesis and degradation of the peptides was assessed by measuring intact, native neuropeptide immunoreactivity in pre-incubated and post-incubated CSF samples, where samples were incubated at 37 degrees C for 1 h. Pre-incubation radioimmunoassay measurements reflected the content of intact peptides present in lumbar CSF at the time of sampling, and post-incubation measurements assayed the amount of peptide that had remained embedded within its precursors [cryptic methionine enkephalin (ME)] and that had been released by the action of CSF peptidases. Significant differences were found in post-incubation samples for the amount of proenkephalin A [ME, leucine enkephalin (LE)] and tachykinin [substance P (SP)] peptides. For example, significant differences were observed for ME-like immunoreactivity (C versus cryptic), SP-like immunoreactivity (PNR versus PR), and LE-like immunoreactivity (PR versus C). No significant differences were observed among the peptides within the pre-incubation samples.(ABSTRACT TRUNCATED AT 250 WORDS)
Harajiri
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Charles B. Stout Neuroscience Mass Spectrometry Laboratory, University of Tennessee, Memphis 38163.
Wood
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Desiderio
D M
DM
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004745:Enkephalins / Q000134:cerebrospinal fluid*; D006801:Humans; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000134:cerebrospinal fluid; D011333:Pro-Opiomelanocortin / Q000134:cerebrospinal fluid*; D011498:Protein Precursors / Q000134:cerebrospinal fluid*; D011863:Radioimmunoassay; D015320:Tachykinins / Q000134:cerebrospinal fluid*
D004745:Enkephalins; D009479:Neuropeptides; D011498:Protein Precursors; D015320:Tachykinins; C029992:proenkephalin; C054618:protachykinin; D011333:Pro-Opiomelanocortin