Effects of protein-modifying reagents on brain tryptamine binding sites: possible involvement of a thiol group in temperature-induced high-affinity [3H]tryptamine binding sites.
To investigate the biochemical nature of temperature-induced high-affinity [3H]tryptamine binding sites, we subjected whole rat brain synaptic membranes to treatment with various protein-modifying reagents and examined the subsequent [3H]tryptamine binding properties of the membranes. Pretreatment of the membrane preparations with NEM, NBS, PCMB, PAPMA and MA, but not with iodoacetamide, DTT, glutathione and cysteine, reduced the [3H]tryptamine binding. In addition, to at least approx. 10(-4) M, the inactivation properties of NEM, PCMB, PAPMA and MA, except for NBS, were temperature-dependent. Furthermore, it was revealed that the Scatchard plot of [3H]tryptamine binding in membranes pretreated with these thiol reagents conformed to a curved line, as well as in the case of the control membranes. Nonlinear regression analysis of these data showed that NEM decreased the Bmax values of both the high and low affinity binding sites with no significant alteration in the KD values, whereas PCMB, PAPMA and MA increased only the KD value of the high affinity sites, accompanying the decrease of the Bmax values of both sites. These results indicate that the temperature-induced high-affinity [3H]tryptamine binding molecule(s) is a thiol protein.
Serikyaku
S
S
Group of Neuropharmacology, Josai University, Saitama, Japan.
Saito
M
M
Ishitani
R
R
D000818:Animals; D001923:Brain Chemistry / Q000187:drug effects*; D066298:In Vitro Techniques; D007202:Indicators and Reagents; D007700:Kinetics; D008297:Male; D018377:Neurotransmitter Agents / Q000378:metabolism; D011506:Proteins / Q000502:physiology*; D051381:Rats; D011919:Rats, Inbred Strains; D011985:Receptors, Serotonin / Q000187:drug effects / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D013570:Synaptic Membranes / Q000187:drug effects / Q000378:metabolism; D013696:Temperature
D007202:Indicators and Reagents; D018377:Neurotransmitter Agents; D011506:Proteins; D011985:Receptors, Serotonin; D013438:Sulfhydryl Compounds