Dimitri E. Grigoriadis

Corticotrophin-releasing factor receptors: from molecular biology to drug design

Corticotrophin-releasing factor (CRF) acts within both the brain and the periphery to coordinate the overall response of the body to stress. The involvement of the CRF systems in a variety of both CNS and peripheral disease states has stimulated great interest in this peptide as a potential site of therapeutic intervention. The recent cloning of multiple CRF receptor subtypes has precipitated a new era in CRF research that has allowed precise molecular, pharmacological and anatomical examination of mammalian CRF receptors. In this article, Derek Chalmers and colleagues highlight the major differences between the two classes of CRF receptors, CRF1 and CRF2, and a functionally related CRF-binding protein, and discuss the relevance of these sites to the ongoing development of CRF-based therapeutics.

Dopamine Transporter: Biochemistry, Pharmacology and Imaging

In recent years, there has been substantial progress in studying the dopamine transporter, a unique component of the functioning dopaminergic nerve terminal. The transporter has been studied by direct binding techniques using a variety of ligands which function as inhibitors of transport. Analogues of these ligands have been used as photoaffinity labels to solubilize and further characterize the transporter. While a variety of drugs bind to the transporter, it is clear that the transporter may serve as an important drug receptor, particularly for the reinforcing properties of some psychostimulants such as cocaine. An extension of the in vitro ligand-binding studies reveals that it is possible to preferentially label the transporter in vivo. The success of in vivo labeling has lead to successful positron emission tomographic scanning studies of the transporter. These studies in turn have revealed the usefulness of imaging the transporter, a measure of the presence of dopaminergic nerve terminals, as a potential diagnostic tool in Parkinsons disease.

Age-related decreases in corticotropin-releasing factor (CRF) receptors in rat brain and anterior pituitary gland

Corticotropin-releasing factor (CRF) receptors were measured in discrete areas of brain and in anterior pituitary of 4-, 12-, 18-, and 24-month-old male Fischer rats. No significant age-related alterations in [125I]ovine CRF binding were observed in the olfactory bulb, cerebral cortex, hippocampus, brainstem, and cerebellum; there was a trend for CRF binding to decrease in the striatum as a consequence of aging. Significant age-related decreases were observed in 125I-ovine CRF binding in the anterior pituitary and hypothalamus with maximal reductions of 60 and 27%, respectively. Saturation analysis in the anterior pituitary indicated an age-related reduction in the density of CRF receptors (i.e. Bmax) without an alteration in the affinity (i.e. Kd) of CRF for its binding site. Northern analysis of proopiomelanocortin (POMC) mRNA in the anterior pituitary indicated no significant differences in the levels of POMC mRNA between 4- and 24-month-old rats. These and other data suggest that the age-related decrease in anterior pituitary CRF receptors may be due to hypersecretion of hypothalamic CRF rather than a loss of corticotropes in the anterior pituitary.

[125]I-Spectramide: A novel benzamide displaying potent and selective effects at the D2 dopamine receptor

The new substituted benzamide Spectramide, (N-[2-[4-iodobenzyl-N-methylamino]-2-methoxy-4-ethyl]-5-chloro- methylamine] benzamide) labelled with 125I was used as a potent and highly selective dopamine-D2 receptor antagonist in rat striatal homogenates for in vitro receptor binding. Kinetic experiments demonstrated the reversibility of the binding and the estimated Kd from saturation analysis was 25 pM, with a Bmax of 20 pmol/g of tissue. Competition studies showed that spectramide did not interact potently with the D1 or dopamine-uptake site. Drugs known to interact with other receptor systems were weak competitors of the binding, while binding was potently inhibited by other D2 antagonists, such as spiperone and eticlopride. These data indicate that Spectramide binds selectively and with high affinity to the dopamine D2 receptors, and may prove to be a useful tool for the study of these receptors in vivo using PET or SPECT.

32 - Biochemical, Pharmacological, and Autoradiographic Methods to Study Corticotropin-Releasing Factor Receptors

Publisher Summary This chapter discusses the biochemical, pharmacological, and autoradiographic methods to study corticotropin-releasing factor (CRF) receptors. CRF, a 41 residue peptide, is the primary physiological regulator or propiomelanocortin-derived peptide secretion from the pituitary gland. In addition to its endocrine effects, there is increasing evidence suggesting that CRF may act as a neurotransmitter or neuromodulator in the central nervous system where it appears to have a major role in coordinating the autonomic, behavioral, and endocrine response of the organism to stress. Nonspecific binding is defined as the binding of a radioligand that remains in the presence of an excess of unlabeled drug known to interact with the binding site. Guanine nucleotides play an integral role in the regulation of receptor-mediated events for many receptor systems. In the absence of antibodies to the receptor proteins themselves, anti-idiotypic antibodies can be raised to compounds known to bind to receptors and used as immunohistochemical agents. The labels can be visualized directly at the light microscopic level and thus, the receptors to which these probes are attached can be visualized directly.

Identification, Autoradiographic Localization, and Modulation of Interleukin 1 Receptors in Brain–Endocrine–Immune Axis: Methodology and Overview

Publisher Summary The cytokine interleukin 1 (IL-1) is one of the key mediators of immunological and pathological responses to stress, infection, and antigenic challenge. In addition to its immune effects, a role has been postulated for IL-1 as a neurotransmitter/neuromodulator/growth factor in the central nervous system (CNS). The central as well as peripheral administration of IL-1 has potent neuroendocrine actions, including the stimulation of the hypothalamic-pituitary-adrenocortical axis and the inhibition of the hypothalamic-pituitary-gonadal axis. These effects of IL-1 are presumably mediated through the actions of the cytokine at specific high-affinity receptors. Recombinant human IL-lα and IL-1β bind to both type I receptors, on T-cells, fibroblasts, keratinocytes, endothelial cells, synovial lining cells, chondrocytes, and hepatocytes, and type II receptors on various B cell lines, including the Raji human B cell lymphoma line. The differential selectivity of IL-1 and IL-lra for type I versus type II IL-1 receptors makes them useful ligands for determining further the characteristics of IL-1 receptors in the brain-endocrine-immune axis.