Cold-restraint stress reduces [3H]etorphine binding to rat brain membranes: influence of acute and chronic morphine and naloxone.

[3H]Etorphine binding was characterized in rat brain homogenates depleted of endogenous opioids from animals acutely and chronically treated with morphine or naloxone and either unstressed or subjected to a 3-h restraint period in the cold. There was significant reduction in the number of high-affinity opiate binding sites in brain tissue from stressed as compared to unstressed animals. Despite the fact that the opiate drug regimens used produce marked behavioral and physiological effects, stress-induced opiate receptor depletion was not influenced by the drugs or by withdrawal. The various drug treatments also failed to produce significant changes in opiate receptor site densities or affinities in either stressed or unstressed animals. We propose that persistent activation of opiate receptors by endogenous opioids released during restraint stress leads to receptor 'down-regulation'.

Effects of morphine and naloxone on stress ulcer formation and gastric acid secretion.

The effect of both acute and chronic morphine and naloxone on restraint-stress gastric ulcer formation and on basal gastric acid secretion were examined in the conscious rat. Acute morphine administration produced a dose-related decrease in stress ulceration and basal gastric acid secretion. Acute naloxone treatment resulted in a dose-related increase in stress ulcer formation and markedly augmented gastric acid secretion. Naloxone (4.0 mg/kg) antagonized the ulcer-reducing effects at all doses of morphine tested (4.0-32.0 mg/kg). Morphine-dependent rats, restrained during spontaneous or naloxone-precipitated withdrawal, exhibited the most severe ulceration. However, only those subjected to naloxone-precipitated withdrawal produced a significant increase in gastric acid output. Chronic treatment with naloxone or with chronic naloxone followed by morphine (16.0 mg/kg) resulted in augmented stress ulcer formation relative to all acutely treated groups. Both chronic naloxone-treated groups exhibited markedly enhanced gastric secretion. These data suggest that central and/or peripheral opiate receptors can modulate both basal and stress-perturbed gastric function.

Apparent competitive inhibition of radioligand binding to receptors: experimental and theoretical considerations in the analysis of equilibrium binding data.

Radioligand binding and displacement experiments are often interpreted in terms of simple competition between two ligands for occupancy of a single binding site on a receptor. Given our current understanding of the complexities of receptor structure and function, it is probable that more complex interactions occur in many cases. By analysis of a hypothetical two-site receptor model, we show that apparent competitive inhibition can arise in several ways, depending on the specificities of the two sites and the interactions between them. We show that binding experiments can in some cases be used to rule out certain models from among a group of apparently plausible ones, provided that experimental criteria are met which permit a meaningful statistical comparison of models to be made. Ideally, these should include: i) an independent study of ligand and inhibitor binding in the absence of each other; ii) carrying out saturation binding and displacement experiments over as wide a range of ligand and inhibitor concentrations as possible; iii) computerized curve-fitting and statistical analysis as a tool for model-testing. While practical limitations may restrict the attainment of such goals, a thorough study of the equilibrium binding properties of a particular receptor system provides the foundation for the design of more definitive experiments at the molecular level, upon which the proof of any binding model ultimately must rest.

ACTH receptors in nervous tissue. High affinity binding-sequestration of [125I]Phe2,Nle4]ACTH 1-24 in homogenates and slices from rat brain.

We have demonstrated specific, high affinity binding of a biologically active Tyr23-monoiodinated derivative of ACTH, [125I][Phe2,Nle4]ACTH 1-24, in rat brain homogenates. Similarly, in metabolically inhibited and noninhibited rat whole brain slices there is a specific "binding-sequestration" process that is dependent on time, protein concentration, and pH. In homogenates, binding curves were best described by a two-site model and provided the following parameters: Kd1 = 0.65 +/- 0.47 nM, Bmax1 = 21 +/- 41 fmol/mg protein; Kd2 = 97 +/- 48 nM, Bmax2 = 3.5 +/- 1.8 pmol/mg protein. In metabolically viable brain slices, concentration-competition curves of [125I][Phe2,Nle4]ACTH 1-24 binding-sequestration can be described by three components (Kd1 = 14 +/- 24 nM, Bmax1 = 50 +/- 95 fmol/mg protein; Kd2 = 2.4 +/- 1.9 microM, Bmax2 = 44 +/- 49 pmol/mg protein; Kd3 = 0.16 +/- 1.0 mM, Bmax3 = 5.3 +/- 54 nmol/mg protein). Metabolic inhibition, by removal of glucose and addition of 100 microM ouabain, abolishes the lowest affinity, highest capacity binding-sequestrian component only (Kd1 = 7.1 +/- 14 nM, Bmax1 = 8.7 +/- 16 fmol/mg protein; Kd2 = 7.4 +/- 4.49 microM, Bmax2 = 37 +/- 27 pmol/mg protein). The two binding-sequestration parameter estimates obtained from metabolically inhibited tissue slices are not significantly different from those of the two higher affinity components obtained with noninhibited tissue. Thus, metabolic inhibition permits demonstration of ACTH receptor binding only, unconfounded by sequestration or internalization of ligand:receptor complexes.(ABSTRACT TRUNCATED AT 250 WORDS)