Binding of 3H-β-endorphin to rat brain membranes: Characterization of opiate properties and interaction with ACTH

doi:10.1016/0014-2999(80)90363-5

European Journal of Pharmacology

Volume 64, Issue 1, 30 May 1980, Pages 1-8

https://doi.org/10.1016/0014-2999(80)90363-5 Get rights and content

Abstract

The binding of tritiated β-endorphin (³H-β-EP) to brain homogenates is described. This had been difficult to achieve due to the lack of availability of ³H-β-EP and to technical difficulties associated with high non-specific binding of β-EP. We now report that ³H-β-EP binding is saturable, stereospecific, has high affinity and is inhibited by sodium. Its dissociation rate is ten-fold longer than that of naloxone. Its regional distribution exhibits interesting differences from naloxone and enkephalin binding. ACTH_1–24 appears to displace it more effectively than it displaces ³H-naloxone. The results are discussed in terms of multiple transmitter systems and the multiple opiate receptor hypothesis.

References (17)

E. Hazum et al.
Interaction of iodinated human [D-Ala²] β-Endorphin with opiate receptors
J. Biol. Chem.
(1979)
R.B. Holman et al.
Simultaneous determination of indole- and catecholamines in small brain regions in the rat using a weak cation exchange resin
Neuroscience
(1976)
E.J. Simon et al.
Opiate receptors and their interactions with agonists and antagonists
Life Sci.
(1975)
L. Terenius et al.
ACTH-like peptides and opiate receptors in the rat brain: structure-activity studies
European J. Pharmacol.
(1975)
F.E. Bloom et al.
Neurons containing β-endorphin in rat brain exist separately from those containing enkephalin: immunocytochemical studies
F.E. Bloom et al.
Endorphins: profound behavioral effects in rats suggest new etiological factors in mental illness
Science
(1976)
A.F. Bradley et al.
Lipotropin: precursor to two biologically active peptides
Biochem. Biophys. Res. Commun.
(1976)
B.M. Cox et al.
Opioid activity of peptides (β-LPH 61–91), derived from β-lipotropin

There are more references available in the full text version of this article.

Cited by (79)

Brain effects of melanocortins
2009, Pharmacological Research
Citation Excerpt :
α-MSH is 600 times more potent than desacetyl-α-MSH in inducing grooming [290,291]. In contrast, non-acetylated α-MSH and structurally related ACTH analogs are more potent than acetylated melanocortins in antagonizing opioid binding to their receptors and in attenuating opiate-mediated analgesia [292,293]. Differences in melanocortin receptor affinity for α-MSH and desacetyl-α-MSH are slight or absent [294,295].
The melanocortins (α, β and γ-melanocyte-stimulating hormones: MSHs; adrenocorticotrophic hormone: ACTH), a family of pro-opiomelanocortin (POMC)-derived peptides having in common the tetrapeptide sequence His-Phe-Arg-Trp, have progressively revealed an incredibly wide range of extra-hormonal effects, so to become one of the most promising source of innovative drugs for many, important and widespread pathological conditions.
The discovery of their effects on some brain functions, independently made by William Ferrari and David De Wied about half a century ago, led to the formulation of the term “neuropeptide” at a time when no demonstration of the actual production of peptide molecules by neurons, in the brain, was still available, and there were no receptors characterized for these molecules.
In the course of the subsequent decades it came out that melanocortins, besides inducing one of the most complex and bizarre behavioural syndromes (excessive grooming, crises of stretchings and yawnings, repeated episodes of spontaneous penile erection and ejaculation, increased sexual receptivity), play a key role in functions of fundamental physiological importance as well as impressive therapeutic effects in different pathological conditions.
If serendipity had been an important determinant in the discovery of the above-mentioned first-noticed extra-hormonal effects of melanocortins, many of the subsequent discoveries in the pharmacology of these peptides (feeding inhibition, shock reversal, role in opiate tolerance/withdrawal, etc.) have been the result of a planned research, aimed at testing the “pro-nociceptive/anti-nociceptive homeostatic system” hypothesis.
The discovery of melanocortin receptors, and the ensuing synthesis of selective ligands with agonist or antagonist activity, is generating completely innovative drugs for the treatment of a potentially very long list of important and widespread pathological conditions: sexual impotence, frigidity, overweight/obesity, anorexia, cachexia, haemorrhagic shock, other forms of shock, myocardial infarction, ischemia/reperfusion-induced brain damage, neuropathic pain, rheumathoid arthritis, inflammatory bowel disease, nerve injury, toxic neuropathies, diabetic neuropathy, etc.
This review recalls the history of these researches and outlines the pharmacology of the extra-hormonal effects of melanocortins which are produced by an action at the brain level (or mainly at the brain level). In our opinion the picture is still incomplete, in spite of being already so incredibly vast and complex. So, for example, several of their effects and preliminary animal data suggest that melanocortins might be of concrete effectiveness in one of the areas of most increasing concern, i.e., that of neurodegenerative diseases.
Roles of acetylation and other post-translational modifications in melanocortin function and interactions with endorphins
2006, Peptides
Citation Excerpt :
α-MSH was far more potent than desacetyl-α-MSH in inducing grooming, and the threshold dose for the latter was 600 times higher than for α-MSH. In contrast, non-acetylated α-MSH and structurally related ACTH analogs are more potent than acetylated melanocortins in attenuating opiate-mediated analgesia and antagonizing opioid receptor binding [3,120]. Also, i.c.v. injection of desacetyl α-MSH selectively increased slow-wave sleep in rats, whereas α-MSH had no effect on any sleep state [17].
Phylogenetic, developmental, anatomic, and stimulus-specific variations in post-translational processing of POMC are well established. For melanocortins, the role of α-N-acetylation and the selective activities of α, β, and γ forms are of special interest. Acetylation may shift the predominant activity of POMC products between endorphinergic and melanocortinergic actions—which are often in opposition. This review addresses: (1) variations in POMC processing; (2) the influence of acetylation on the functional activity of α-MSH; (3) state- and stimulus-dependent effects on the proportional distribution of forms of melanocortins and endorphins; (4) divergent effects of α-MSH and β-endorphin administration; (5) potential roles of β- and γ-MSH.
No evidence for G-protein-coupled epsilon receptor in the brain of triple opioid receptor knockout mouse
2004, European Journal of Pharmacology
Citation Excerpt :
Interestingly the existence of epsilon sites was suggested in the rat brain also, on the basis of binding studies. [ 3H]β-Endorphin labeling showed a unique regional distribution (Akil et al., 1980; Goodman et al., 1983; Johnson et al., 1982) and distinct sensitivity to bivalent cations (Law et al., 1979) as compared to other tritiated opioid ligands. Later, studies aiming at characterizing the so-called “benzomorphan sites” in rat brain revealed strong similarities with properties of the epsilon receptor described in the rat vas deferens.
Pharmacological approaches have defined the epsilon receptor as a β-endorphin-preferring opioid receptor, described in rat vas deferens and in brain of several species. Only three opioid receptors—mu, delta and kappa—have been cloned and the existence of this additional subtype as a distinct protein remains controversial. Recently, the mouse brain epsilon receptor was detected in a G protein activation assay, as mediating residual β-endorphin activity following pharmacological blockade of mu, delta and kappa receptors. To clarify whether this site is independent from mu, delta and kappa receptors, we performed β-endorphin-induced [³⁵S]GTPγS binding using mice lacking these three receptors (triple knockout mice). We tested both pons–medulla and whole brain preparations. β-Endorphin strongly stimulated [³⁵S]GTPγS binding in wild-type membranes but had no detectable effect in membranes from triple knockout mice. We conclude that the brain epsilon site involves mu, delta and/or kappa receptors, possibly coupled to nonclassical G proteins.
The effect of melanotropic peptides on binding of [<sup>3</sup>H]dihydroalprenolol-hydrochloride to hypothalamic membranes
2001, Peptides
In the present work we studied the interaction of α-melanocyte-stimulating hormone (α-MSH) and ACTH-(1–24) with β-adrenergic receptors in hypothalamic membranes from rat brain. Saturation curves for [³H]dihydroalprenolol-hydrochloride ([³H]DHA) binding in the presence of the peptides revealed a decreased binding capacity (Bmax). The dissociation constant (Kd) was, however, not affected by α-MSH or ACTH-(1–24). These data indicate a non competitive interaction between these melanocortin peptides and [³H]DHA on β-adrenergic receptors in hypothalamic membranes.
α-Melanotropin hormone inhibits the binding of [<sup>3</sup>H]SCH 23390 to the dopamine D<inf>1</inf> receptor in vitro
1998, European Journal of Pharmacology
We have previously demonstrated that the simultaneous presence of α-melanocyte stimulating hormone (α-MSH) and dopamine resulted in a reduction in cyclic AMP (cAMP) levels in slices containing caudate putamen and accumbens nuclei as compared to those treated only with dopamine or α-MSH. This study was carried out to explore if the interaction between α-MSH and dopamine could be explained on the basis of a direct interaction between α-MSH and the dopamine D₁ receptor. Saturation curves for [n-methyl- $^{3} H$ ](R)-(+)-8 chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3-benzazepin-7-ol hemimaleate ([ $^{3} H$ ]SCH 23390) binding in the presence of increasing concentrations of α-MSH were performed. Nonlinear regression in the presence of α-MSH revealed an increased dissociation constant (K_d). The binding capacity (B_max) was not affected by the peptide. These data suggest an apparent competitive interaction between α-MSH and [ $^{3} H$ ]SCH 23390 in striatal membranes on the dopamine D₁ receptor; (K_i=1.2×10⁻⁷ M). The present data show that α-MSH could interact with the dopamine D₁ receptor modulating allosterically the affinity of [ $^{3} H$ ]SCH 23390 for the receptor or by causing a change in the lipid environment of the dopamine receptor, resulting in an inhibition of the ligand binding to it.
Evidence for the existence of the β-endorphin-sensitive 'ε-opioid receptor' in the brain: The mechanisms of ε-mediated antinociception
1998, Japanese Journal of Pharmacology
Recently, μ-, δ- and κ-opioid receptors have been cloned and relatively well-characterized. In addition to three major opioid receptor types, more extensive studies have suggested the possible existence of other opioid receptor types that can be classified as non-μ, non-δ and non-κ. Based upon anatomical and binding studies in the brain, the sensitive site for an endogenous opioid peptide, β-endorphin, has been postulated to account for the unique characteristics of the opioid receptor defined as a putative ε-opioid receptor. Many ε-opioid receptors are functionally coupled to G-proteins. The functional ε-opioid receptors in the brain are stimulated by bremazocine and etorphine as well as β-endorphin, but not by selective μ-, δ- or κ-opioid receptor agonists. ε-Opioid receptor agonists injected into the brain produce profound antinociception. The brain sites most sensitive to ε-agonist-induced antinociception are located in the caudal medial medulla such as the nucleus raphe obscures, nucleus raphe pallidus and the adjacent midline reticular formation. The stimulation of ε-opioid receptors in the brain facilitates the descending enkephalinergic pathway, which probably originates from the brainstem terminating at the spinal cord. The endogenous opioid Met-enkephalin, released in the spinal cord by activation of supraspinal ε-opioid receptors, stimulates spinal δ2-opioid receptors for the production of antinociception. It is noteworthy that the ε-opioid receptor-mediated pain control system is different from that of other opioid systems. Although there appears to be no ε-selective ligand currently available, these findings provide strong evidence for the existence of the putative ε-opioid receptor and its unique function in the brain.

View all citing articles on Scopus

View full text

Binding of 3H-β-endorphin to rat brain membranes: Characterization of opiate properties and interaction with ACTH

Abstract

J. Biol. Chem.

Neuroscience

Life Sci.

European J. Pharmacol.

Neurons containing β-endorphin in rat brain exist separately from those containing enkephalin: immunocytochemical studies

Endorphins: profound behavioral effects in rats suggest new etiological factors in mental illness

Science

Lipotropin: precursor to two biologically active peptides

Biochem. Biophys. Res. Commun.

Opioid activity of peptides (β-LPH 61–91), derived from β-lipotropin

Binding of ³H-β-endorphin to rat brain membranes: Characterization of opiate properties and interaction with ACTH