Pharmacological Characterization of the D1- and D2-Like Dopamine Receptors from the Brain of the Leopard Frog, <i>Rana pipiens</i>

Abstract

The pharmacological profiles of D1- and D2-like dopamine receptors were investigated for native brain receptors in the leopard frog, Rana pipiens, using direct binding assays, which characterize functional receptors rather than assess total receptor protein. We used homogenate assays of R. pipiens fore- and midbrains to determine, via saturation isotherms, that the dissociation constant, Kd, for ³H-SCH-23390 binding to the D1-like receptors was 0.29 nM, and the maximal receptor density, Bmax, was 40 fmoles/mg protein. This compares with the more than 10-fold higher density of D1 sites in rat striatum. Specific binding for the D2-like receptors was measurable using these methods with ³H-spiperone as the ligand. However, saturation of binding was not achieved. This contrasts with the > 400 fmoles/mg protein Bmax in rat striatum. Pharmacological profiles (rank order of potency of displacing drugs) for each receptor type were determined. We used non-radioactive SCH-23390, SKF-38393, sulpiride, and spiperone to displace ³H-SCH-23390 and ³H-spiperone at D1 and D2 receptors, respectively. Parallel displacement assays were performed with rat striatal controls. Results indicated that the relative rank order displacements in anuran dopamine receptors were characteristic of D1- and D2-like receptors. However, the rank orders were not identical to those in mammals. The rank order for affinity at D1-like receptors in both rats and frogs was SCH-23390 > SKF-38393 > spiperone > sulpiride. The rank order for affinity at D2-like receptors was spiperone > SCH-23390 > sulpiride > SKF-38393 in frogs, and spiperone > sulpiride > SCH-23390 > SKF-38393 in rats. SKF-38393 and spiperone had similar affinities for the ‘D1’ receptors in both species. SCH-23390 had a slightly lower affinity for the D1-like receptors in Rana, whereas sulpiride had a significantly lower affinity for Rana D1-like receptors compared to rat D1 receptors. In Rana D2-like receptors, spiperone and sulpiride were significantly less potent compared to rat. However, SCH-23390 and SKF-38393 were equally potent for the D2-like receptors in both species. The results indicate that amphibian brain dopamine receptors fall into two classes similar to the mammalian D1 and D2 subfamilies, but with binding characteristics slightly different from those typically described in mammals. This work represents the first pharmacological characterization of native brain dopaminergic receptors in an anuran amphibian. Because direct binding assays measure the initial aspect of the functional interaction between transmitter and receptor, these data provide an important complement to studies using cell expression systems.

Effects of Amphetamine on Conditioned Place Preference and Locomotion in the Male Green Tree Frog, <i>Hyla cinerea</i>

“…The drug doses used in this study are consistent with studies in other amphibians showing effective doses higher than those used in rodents models [Marler et al, 1995;Glagow and Ewert, 1999;Ten Eyck, 2008]. This increase in effective dose may account for physiological differences between amphibians and mammals, such as differences in metabolic rate (ectotherms vs. endotherms) or differences in receptor pharmacology [Chu et al, 2001]. Although a pharmacological characterization of AMPH has not been examined in amphibians, many mechanisms of dopamine pharmacology show a strong conservation of function.…”

“…Such conservation of function include the neurotoxic effects of MPTP+ [Barbeau et al, 1985] and 6-hydroxydopamine [Endepols et al, 2004] on dopamine neurons in frog CNS, as well as behavioral effects of the mixed dopamine agonist apomorphine on motor behavior [Glagow and Ewert, 1999;Chu and Wilczynski, 2007;Chu and Rankine, 2008]. Dopamine D1-and D2-like receptors have been pharmacologically characterized in the brain of the northern leopard frog and have been shown to possess binding profiles for various dopaminergic ligands that are similar but not as potent as those of the rat striatum [Chu et al, 2001].…”

7α-Hydroxypregnenolone acts as a neuronal activator to stimulate locomotor activity of breeding newts by means of the dopaminergic system

“…It has been reported that in amphibians, dopamine neurons are localized in the medial brain region, such as the posterior tuberal nucleus and ventral tegmental area, and that they project to the rostral brain region, such as the striatum (32,33). It also has been reported that, as in mammal brain, the amphibian brain possesses dopamine D 1 -and D 2 -like receptors (34) and that both haloperidol and sulpiride abolish dopamine actions in amphibians (35). The present and previous studies with amphibians suggest that the stimulatory effect of 7␣-hydroxypregnenolone on locomotor activity is mediated by the dopaminergic system.…”

Sex differences and hormone influences on tyrosine hydroxylase immunoreactive cells in the leopard frog

“…From rostral to caudal, these include a set of periventricular neurons in the preoptic area (POA), a larger population contained within the suprachiasmatic nucleus (SCN), and the most caudal and extensive cell group that occupies the posterior tuberculum (TP) rostrally before moving ventrolaterally into the dorsal hypothalamus (Hy) at the caudal end of the diencephalon. Immunohistochemical studies (Gonzá-lez and Smeets, 1991;González et al 1993;Smeets and Reiner 1995) have shown that these TH-ir cells are dopaminergic, and pharmacological studies by Chu et al (2001) indicate that the forebrain contains both D1-like and D2-like dopamine receptors. Dopaminergic fibers from these cells extend throughout the forebrain and midbrain.…”