Effect of Tris, HEPES, and TES buffers on binding at μ-, δ-, and κ-opioid sites in guinea pig brain

In homogenates of guinea-pig brain minus cerebellum, the δ-binding of 1.5 nM [3H]-[D-Pen2,D-Pen5]enkephalin is little affected by the type and concentration of the three buffers, Tris-HCl, HEPES-KOH, or TES-KOH (10–75 mM). However, the μ-binding of 1 nM [3H][D]A[a2,MePhe4,GIy-ol5]enkephalin or the κ-binding of 1.5 nM [3H]-U-69,593 is influenced by the choice of buffer. A suitable concentration of buffer for further analyses of opioid binding has been found to be 10 mM. At each site, the effects of MgCl2 on binding are the same whether 10 mM Tris-HCl, HEPES-KOH, or TES-KOH are used but variations of the effects of NaCl confirm the view that μ- and κ-sites, but not δ-sites, are affected by choice of buffer. Furthermore, under some assay conditions the effects of NaCl and MgCl2 at the κ-sites of guinea-pig cerebellum differ from their effects in brain minus cerebellum, indicating that these are differences of binding characteristics at the κ-sites of these tissues.