Octopamine receptors in the honey bee and locust nervous system: pharmacological similarities between homologous receptors of distantly related species.

Honey bees are perhaps the most versatile models to study the cellular and pharmacological basis underlying behaviours ranging from learning and memory to sociobiology. For both aspects octopamine (OA) is known to play a vital role. The neuronal octopamine receptor of the honey bee shares pharmacological similarities with the neuronal octopamine receptor of the locust. Both, agonists and antagonists known to have high affinities for the locust neuronal octopamine receptor have also high affinities for the bee neuronal octopamine receptor. The distribution of receptors is more or less congruent between locusts and bees. Optic lobes and especially the mushroom bodies are areas of greatest octopamine receptor expression in both species, which mirrors the physiological significance of octopamine in the insect nervous system. The neuronal octopamine receptor of insects served as a model to study the pharmacological similarity of homologous receptors from distantly related species, because bees and locusts are separated by at least 330 million years of evolution.

The pharmacology of a dopamine receptor in the locust nervous tissue.

A dopamine receptor in the nervous tissue of the desert locust (Schistocerca gregaria Forskâl) was studied using ¿3Hlysergic acid diethylamide (LSD) as the radioligand. Its expression is almost entirely restricted to the mushroom bodies, centres for learning and memory in the insect brain. This G-protein coupled receptor is present in relatively low concentrations in the locust brain (35 fmol/mg protein). The pharmacological characterisation reveals high affinity for the putative natural agonist dopamine (K(i)=28 nM). Substances with high subtype specificity for vertebrate dopamine receptors such as SCH 23390 (K(i)=639 nM) and sulpiride (K(i)=21,200 nM) have low affinity for the locust neuronal dopamine receptor. In opposite, substances with a broad pharmacological profile such as LSD, spiperone (K(i)=7.26 nM), and chlorpromazine (K(i)=9.52 nM) have high affinity properties. Comparison of the pharmacological data reveals no significant homology to any vertebrate dopamine receptor class characterised so far. This uncertainty about the pharmacological relatedness of insect dopamine receptors mirrors the available molecular data. It is almost impossible to classify cloned insect dopamine receptors into vertebrate dopamine receptor schemes. This lack of pharmacological relatedness opens the opportunity to develop highly specific insecticides against insect dopamine receptors.

Voltage-activated whole-cell K+ currents in lamina cells of the desert locust schistocerca gregaria

Voltage-dependent outward currents were studied in freshly dissociated somata of locust lamina cells. These currents were recorded in 142 somata using the whole-cell patch-clamp technique. By measuring the reversal potential at altered external [K+] and by replacing internal K+ with Cs+, we determined that the outward currents were carried by K+. The outward currents consist of a transient A-type K+ current (KA) and a delayed-rectifier-like K+ current (KD). Amongst the cells studied, we observed two distinct groups of cells. The most obvious difference between the two groups is that in group I cells the total outward current is dominated by KA (KA/KD=12.5), whereas in group II cells KA makes a smaller contribution (KA/KD=2.1). Furthermore, in cells of group I, the KA current shows a steeper voltage-dependence of activation, where VG50 is -29.9 mV and s is 11.9 (N=22), and inactivation, where VI50 is -84.5 mV and s is -6.3 (N=18), compared with the KA current in cells of group II: VG50=-7.9 mV; s=26.6 (N=36) and VI50=-68.4 mV; s=-7.5 (N=21) (VG50 is the voltage at which the whole-cell conductance G is half-maximally activated, VI50 is the voltage of half-maximal inactivation and s is the slope of the voltage-dependence). The transient KA current in group I cells decayed mono-exponentially. The decay of the KA current in group II cells was fitted with a double-exponential curve and was significantly faster than in group I cells. In contrast to the large differences in KA currents, the KD currents appeared to be quite similar in the two groups of cells.

Epinastine, a highly specific antagonist of insect neuronal octopamine receptors.

The tetracyclic compound epinastine (3-amino-9, 13b-dihydro-1H-dibenz(c,f)imidazo(1,5a)azepine hydrochloride) that was recently introduced as a vertebrate histamine H1 receptor antagonist has also high affinity for insect neuronal octopamine receptors. This holds true for the neuronal octopamine receptor from the locust (Ki = 2 Nm) as well as from the honey bee nervous system (Ki = 1.1 Nm). In addition to its high affinity, it has a high degree of specificity. Its affinity for other insect receptors for biogenic amines, such as 5-hydroxytryptamine, dopamine, histamine, and tyramine, is at least four orders of magnitude lower. Therefore, epinastine could serve as a highly specific antagonist of octopamine receptors that enables physiological dissection of octopaminergic neurotransmission within the nervous system of insects. To demonstrate these abilities, epinastine was used to inhibit the visually evoked activity of an identified interneuron in the visual pathway which is known to be modulated by octopamine.

Pharmacological characterization of a 5-HT receptor in locust nervous tissue.

A 5-HT receptor in the nervous tissue of the desert locust (Schistocerca gregaria Forsk.) was investigated, using [3H]LSD (lysergic acid diethylamide) as the radioligand. [3H]LSD labels in addition a putative dopamine receptor whose specific [3H]LSD binding nevertheless could easily be diminished by co-incubation with 1 microM dopamine. The binding site was characterized by a KD of 1.64 nM, and a maximal concentration of binding sites of 79.8 fmol/mg protein. Pharmacological investigation revealed a relatively low affinity for the putative natural agonist, serotonin (KI = 0.209 microM). In contrast to the high affinity of classical serotonergic antagonists (e.g. dihydroergotamine or (+)-butaclamol) substances with subtype specificity such as 8-OH-DPAT (8-hydroxyl-1-(N,N-dipropyl)-aminotetralin) or ketanserin have only moderate affinities. Quantitative comparison of the pharmacological data demonstrated that there is obviously no pharmacological homology with vertebrate 5-HT receptors characterized so far. The only receptors with a close pharmacological relationship to the 5-HT receptor of locusts are the 5-HTdro1 receptor expressed in Drosophila nervous tissue and a 5-HT receptor in snail nervous tissue which might be homologous to that of locusts. The 5-HT receptor investigated, was shown to be G-protein-coupled, as addition of stable GTP analogues or depletion of Mg2+ ions from the incubation medium led to agonist-specific lowering of the affinity.

Octopamine receptors in locust nervous tissue.

The octopamine binding site in the nervous tissue of the migratory locust Locusta migratoria is identified as an octopamine receptor of class 2. The binding of octopamine to the binding site is saturable, reversible, stereospecific and shows a pharmacological profile typical for octopamine receptors. Saturation analysis results in a single class of non-interacting binding sites (KD = 7.9 +/- 0.9 nM; Bmax = 160 fmol/mg). The pharmacological analysis shows that the phenyliminoimidazolidines NC7 and NC5 (Ki = 0.29 and 0.87 nM, respectively) are the most potent agonists, and that mianserin (Ki = 1.20 nM) is the most potent antagonist ever reported for octopamine receptors in direct binding studies.