Tyramine receptor & Tyramine receptorII: Biological Overview | References
Gene name - Tyramine receptor & Tyramine receptorII
Cytological map position - 90C2-90C3 & 90C2-90C2
Function - G-protein coupled receptor
Symbol - TyrR & TyrRII
Classification - 7 transmembrane receptor (rhodopsin family)
Cellular location - surface transmembrane
Neuromodulators influence the activities of collections of neurons and have profound impacts on animal behavior. Male courtship drive is complex and subject to neuromodulatory control. Using the fruit fly Drosophila melanogaster, this study identified neurons in the brain (inferior posterior slope; IPS) that impact courtship drive and are controlled by tyramine-a biogenic amine related to dopamine, whose roles in most animals are enigmatic. A tyramine-specific receptor, TyrR, which is expressed in IPS neurons, was knocked out. Loss of TyrR leads to a striking elevation in courtship activity between males. This effect occurs only in the absence of females, as TyrRGal4 mutant males exhibit a wild-type preference for females. Artificial hyperactivation of IPS neurons causes a large increase in male-male courtship, whereas suppression of IPS activity decreases male-female courtship. The study concludes that TyrR is a receptor for tyramine, and suggests that it serves to curb high levels of courtship activity through functioning as an inhibitory neuromodulator (Huang, 2016).
Neurotransmitters and neuromodulators that are derived from tyrosine are evolutionarily conserved, and are critical mediators of animal behavior. Dopamine and the related catecholamine norepinephrine are synthesized through a simple pathway that begins with conversion of tyrosine into dihydroxyphenylalanine (DOPA). Tyrosine is also a substrate for production of octopamine, which is structurally similar to norepinephrine. Octopamine is produced in both mammals and invertebrates, although its role as a neuromodulator and neurotransmitter is best characterized in insects, where it promotes an array of behaviors. These range from male aggression to learning and memory in flies, female post-mating behaviors, sleep, foraging, and others (Huang, 2016).
The biosynthesis of octopamine is initiated by decarboxylation of tyrosine to produce tyramine, which is present at low levels in many mammalian tissues, including the brain. Due to its concentration in trace amounts, it has long been thought to serve primarily as a biosynthetic precursor of octopamine, and not as a neuroactive chemical in its own right. Nevertheless, the discovery of a specific family of G protein-coupled receptors (GPCRs), some members of which are activated primarily by tyramine, raises the possibility that tyramine may function independently as a neuromodulator. Indeed, the concentration of tyramine is altered in a variety of human neurological disorders, including schizophrenia, Parkinson's disease, attention deficit hyperactivity disorder, Tourette syndrome, and phenylketonuria. Nevertheless, the functions of tyramine are enigmatic, especially in mammals (Huang, 2016).
The brains of the fruit fly harbor populations of neurons that produce tyramine, and not octopamine, arguing against a trivial role for tyramine exclusively as a metabolic intermediate. A few experiments in insects address this possibility. For example, a Drosophila mutation affecting a receptor for both octopamine and tyramine (Oct-TyrR) results in reduced odor avoidance (Kutsukake, 2000). However, it is unclear whether the phenotype reflects a role for octopamine or tyramine, because Oct-TyrR is activated by tyramine and octopamine with similar potency. Application of tyramine to Drosophila tissue, or injections of tyramine into the blowfly or moth, produces a variety of physiological responses. However, the tyramine might be metabolically converted to other biogenic amines that elicit function. At this time, there is no clear genetic evidence indicating a role for tyramine as an independent neuromodulator in Drosophila. Despite the presence of tyramine in the brains of animals that include mammals and insects, Caenorhabditis elegans is the only organism for which genetic evidence supports a role of tyramine as a neuromodulator (Huang, 2016).
The Drosophila genome encodes multiple GPCRs that are activated by biogenic amines, one of which (TyrR) is activated specifically by tyramine, but not by the other biogenic amines tested, including octopamine, dopamine, serotonin, and histamine. This study has generated a null mutation in TyrR and found that the mutant males displayed a profound increase in male-male courtship but no change in gender preference. TyrR was expressed and functioned in a set of tyramine-responsive neurons in the Drosophila brain called the inferior posterior slope (IPS). Genetic hyperactivation of IPS neurons induced a significant elevation in male-male courtship, similar to the mutant males. Conversely, inactivation of these neurons decreased male-female courtship. It is concluded that basal IPS activity is required to permit sufficient levels of sexual drive for male-female courtship. In addition, through activation of TyrR, it is suggested that tyramine serves as an inhibitory neuromodulator to reduce sexual drive (Huang, 2016).
Nearly all wild-type Drosophila males court and mate with females. However, among wild-type flies, the frequency of male-male courtship is low. Nevertheless, there are multiple mutations that increase male-male courtship. The changes in behavior are typically due to deficits in identifying males, such as occurs upon elimination of male pheromones or the corresponding receptors. This study found that TyrRGal4 mutant males exhibit a dramatic increase in male-male sexual activity. In contrast to previous mutations that increase male-male courtship, TyrRGal4 flies discriminate between male and females. When provided a choice between the two genders, the TyrRGal4 mutants select females at the same high proportion as wild-type males. These results suggest that the strong male-male courtship activity was not due to a deficit in sensing repulsive male pheromones. Furthermore, TyrRGal4 males also exhibited increased courtship toward young and aged females. These phenotypes were due to loss of TyrR and not potential effects of the mini-white transgene, because the TyrR phenotype with a wild-type TyrR transgene. Moreover, heterozygous control males harboring the mini-white gene (TyrR/+) display wild-type levels of courtship behavior. Furthermore, the increased male-male courtship was recapitulated by RNAi knockdown of TyrR. Consistent with the conclusion that tyramine modulates male courtship activity, Tdc2, but not Tβh, mutant males show elevated levels of male-male courtship (Huang, 2016).
It is proposed that the TyrR-expressing neurons control overall male sexual drive. In support of this concept, suppressing the normal activity of TyrR-expressing neurons in wild-type males significantly reduced male-female courtship behavior. This manipulation slightly reduced male-male courtship behavior in wild-type. However, the effect was not statistically significant, because basal male-male courtship activity was very low. Nevertheless, silencing TyrR+ neurons in TyrRGal4 mutant males eliminated the high male-male courtship activity. Conversely, when the TyrR+ neurons were artificially activated in wild-type males, the animals displayed a strong elevation in male-male courtship behavior. Thus, the dramatic increase in male-male courtship reflected an increase in overall sexual activity, rather than an increase in same-sex preference. Based on GRASP studies, it is suggested that the TyrR+ neurons function through the FruM neural circuits. Thus, the normal low activity of TyrR-positive neurons is permissive for male-female courtship. Higher activity stimulates greater courtship drive such that the animals will also court males, but only if females are not present, because even at artificially elevated levels of activity the males still prefer females if both gender targets are available. This role for TyrR-expressing neurons differs from P1 neurons, which promote distinct behaviors, aggression and courtship, at low and high activity levels, respectively (Huang, 2016).
The TyrRGal4 phenotype was due to a requirement for tyramine for controlling courtship behavior, because this study found that TyrR-expressing neurons were activated by tyramine, but not octopamine, consistent with in vitro data indicating that TyrR is activated specifically by tyramine (Cazzamali, 2005). Tyramine is most likely acting as a neuromodulator, rather than as a neurotransmitter, because TyrR is a GPCR rather than an ionotropic receptor. In further support of this model, no GRASP signals were detected using the Tdc2-LexA and TyrRGal4, suggesting that the tyramine-producing neurons are not in direct contact with the TyrR-expressing neurons. However, a caveat is that the Tdc2-LexA recapitulates only a subset of the Tdc2 neurons (Huang, 2016).
A neuromodulator can either be excitatory or inhibitory, depending on the receptor that is activated. The data in this study suggest that as a consequence of activating TyrR, tyramine serves as an inhibitory rather than excitatory neuromodulator, which curbs sexual activity. In favor of this proposal are the genetic activation and inactivation experiments. Artificial stimulation of TyrR-expressing neurons increased male-male courtship, whereas inhibition of these neurons reduced male-female courtship (Huang, 2016).
The model that TyrR is an inhibitory neuromodulator receptor in vivo is consistent with in vitro studies showing that tyramine reduces the amplitude of excitatory junction potential (EJP) in neuromuscular junctions (Ormerod, 2013; Nagaya, 2002). In vivo Ca2+ imaging results, as well as an in vitro analysis, indicate that TyrR is coupled to Gq, which typically leads to neuronal activation. However, loss of Gq/G11 signaling can increase neuronal activity as well. This could potentially occur through inhibiting glutamate release, gating of a Ca2+ -activated K+ channel, or promiscuous coupling of Gq/G11 -coupled receptors to Gi/o G proteins. In support of this latter possibility, two related Drosophila catecholamine receptors are coupled to both Gq and Gi proteins (Huang, 2016).
This study found that TyrR activity was required in a small group of neurons (TyrRIPS) in the brain for controlling courtship drive. Tyramine-induced inhibition of TyrRIPS neurons was strictly dependent on TyrR, because the response was eliminated in TyrRGal4 mutant brains. Based on findings using the GRASP technique, it is proposed that TyrRIPS neurons may form synaptic connections with FruM neurons, which regulate courtship. It is proposed that courtship behavior is enhanced by release of acetylcholine from basal or highly activated TyrRIPS neurons. In support of this proposal, TyrRIPS cells expressed ChAT, and knockdown of ChAT in these cells reduced courtship behavior (Huang, 2016).
In conclusion, the findings show that in Drosophila, tyramine is not simply a biosynthetic intermediate for octopamine. Rather, it has an important function in the neuromodulation of male courtship drive through its specific receptor, TyrR. However, it does not affect gender preference. Given the presence of tyramine as a trace monogenic amine in the mammalian brain, the question arises as to whether tyramine also functions in mammals as a neuromodulator of behavior (Huang, 2016).
The monoamines octopamine and tyramine, which are the invertebrate counterparts of epinephrine and norepinephrine, transmit their action through sets of G protein-coupled receptors. Four different octopamine receptors (Oamb, Octβ1R, Octβ2R, Octβ3R) and 3 different tyramine receptors (TyrR, TyrRII, TyrRIII) are present in the fruit fly Drosophila melanogaster. Utilizing the presumptive promoter regions of all 7 octopamine and tyramine receptors, the Gal4/UAS system is utilized to elucidate their complete expression pattern in larvae as well as in adult flies. All these receptors show strong expression in the nervous system but their exact expression patterns vary substantially. Common to all octopamine and tyramine receptors is their expression in mushroom bodies, centers for learning and memory in insects. Outside the central nervous system, the differences in the expression patterns are more conspicuous. However, four of them are present in the tracheal system, where they show different regional preferences within this organ. On the other hand, TyrR appears to be the only receptor present in the heart muscles and TyrRII the only one expressed in oenocytes. Skeletal muscles express octβ2R, Oamb and TyrRIII, with octβ2R being present in almost all larval muscles. Taken together, this study provides comprehensive information about the sites of expression of all octopamine and tyramine receptors in the fruit fly, thus facilitating future research in the field (El-Kholy, 2015).
In invertebrates, the phenolamines, tyramine and octopamine, mediate many functional roles usually associated with the catecholamines, noradrenaline and adrenaline, in vertebrates. The α- and β-adrenergic classes of insect octopamine receptor are better activated by octopamine than tyramine. Similarly, the Tyramine 1 subgroup of receptors (or Octopamine/Tyramine receptors) are better activated by tyramine than octopamine. However, recently, a new Tyramine 2 subgroup of receptors was identified, which appears to be activated highly preferentially by tyramine. This study examined immunocytochemically the ability of CG7431 (TyrR), the founding member of this subgroup from Drosophila melanogaster, to be internalized in transfected Chinese hamster ovary (CHO) cells by different agonists. It was only internalized after activation by tyramine. Conversely, the structurally related receptor, CG16766 (TyrRII), was internalized by a number of biogenic amines, including octopamine, dopamine, noradrenaline, adrenaline, which also were able to elevate cyclic AMP levels. Studies with synthetic agonists and antagonists confirm that CG16766 has a different pharmacological profile to that of CG7431. Species orthologues of CG16766 were only found in Drosophila species, whereas orthologues of CG7431 could be identified in the genomes of a number of insect species. It is proposed that CG16766 represents a new group of tyramine receptors, which we have designated the Tyramine 3 receptors (Bayliss, 2013).
The two putative Drosophila tyramine receptors, CG7431 and CG16766, lie adjacent to each other on the right arm of Chromosome 3 and have a very similar intron-exon structure with five coding exons. They share a 61% sequence identity and a 79% sequence similarity. They are much more similar to each other than to the Drosophila Oct/Tyr receptor (CG7485 or DmTAR1) (CG7431: 30% Identical and 47% Similar; CG16766: 44% Identical and 65% Similar). Thus, they are likely to be paralogue genes that have arisen by gene duplication. The results of this study suggest that the two genes are likely to have evolved different functions over the course of evolution (Bayliss, 2013).
CG7431 is a highly unusual insect biogenic amine receptor in that it shows a very high specificity for the biogenic amine tyramine in calcium-release assays when expressed in CHO cells (Cazzamali, 2005). Structurally related biogenic amines, such as octopamine, synephrine and dopamine did not show any effects up to a concentration of 100 μM on CG7431 (Cazzamali, 2005) or on its species homologue from Bombyx (BmTAR2). It was asked if the Drosophila receptor might show agonist-specific coupling for different biogenic amines if it was assayed using different second messenger systems. However, neither tyramine nor any of the other agonists tested showed any effects in cyclic AMP assays up to a concentration of 1 μM. To attempt to determine if the receptor might be able to couple to some other untested second messenger pathway, a series of internalization studies was carried out to see if the receptor could be activated by biogenic amines other than tyramine. A considerable number of biogenic amine-activated GPCRs show agonist-dependent internalization. However, in these studies, tyramine was the only biogenic amine to internalize the receptor from the plasma membrane to a perinuclear localization. As another Drosophila GPCR, CG 18314 or DmDopEcR, also shows a very high specificity for dopamine in cyclic AMP stimulation assays, but also responds to insect steroids such as ponasterone A, ecdysone and 20-hydroxy-ecdysone, this study explored the possibility that CG7431 might also be a steroid receptor. However, ecdysone, 20-hydroxy-ecdysone or 17β-estradiol were not able to induce the internalization of the receptor. Furthermore, as tyrosine-containing neuropeptides effectively contain a 'tyramine motif', the effectiveness of neuropeptides including, proctolin and Leu-enkephalin, were tested for their ability to internalize CG7431. However, neither of the peptides was effective. Thus, CG7431 is either a highly unusual biogenic amine receptor with a very high specificity for tyramine or has a hitherto unknown endogenous agonist (Bayliss, 2013).
CG16766 showed a very different pharmacology to CG7431 in terms of both its agonist-mediated internalization and biogenic amine specificity. In both cases, several other biogenic amines were also effective, similar to the Drosophila Oct/Tyr receptor. Thus, tyramine and phenylethylamine were almost equally effective on the receptor, and their effects were reduced by adding a β-hydroxyl group and an N-terminal methyl group. Interestingly, phenylethylamine was also able to block the binding of [3H] tyramine to the BmTAR2 receptor, but was not able to activate it. In terms of synthetic agonists, CG16766 showed some similarities with the locust Tyr 1 receptor, the Bombyx Tyr 2 receptor and the Drosophila β-adrenergic octopamine receptors in that it was activated by the α-adrenergic agonists, naphazoline and tolazoline. In contrast to other insect octopamine and tyramine receptors, synthetic antagonists such as yohimbine, mianserin and chlorpromazine were not able to block the actions of tyramine on CG16766 (Bayliss, 2013).
CG7431 and CG16766 also differ in their reported tissue expression patterns in Drosophila (Chintapalli, 2007), further suggesting that they may undertake different physiological roles in the animal. The expression of CG7431 is enriched in the brain, thoracicoabdominal ganglion and in the midgut in the adult and in the CNS, tubule and hindgut in larvae. This suggests that it might have functional roles as a neurotransmitter or neuromodulatory receptor in the nervous system and in the control of gut function. The orthologous receptor BmTAR2 was detected in the brain and nerve cord of the fifth instar larvae, but not in the silk gland, midgut or Malpighian tubules. The expression of the CG16766 receptor, on the other hand, is highly enriched in the crop and eye in adults and in the tubule and hindgut in larvae. This suggests that it might have a specific role in the modulation of vision and in the storage of food. These data further suggest that the paralogous receptors may have gained different functions during the course of evolution (Bayliss, 2013).
A phylogenetic analysis of the Drosophila melanogaster CG7431 and CG16766 receptor sequences clearly shows that both receptor sequences group together with orthologous sequences from the other Drosophila species of the subgenus Sophophora, such as those of D. simulans, D. sechellia, D. yakuba and D erecta. In addition, the CG7431 sequence also clusters with the Tyramine 2 receptors from a number of other insect species. However, it was not possible to identify any homologous sequences to that of CG16766 in any of the sequenced insect genomes, confirming and extending the observation of Huang (2009), who could not find a CG16766-like sequence in the Bombyx genome. Thus, CG16766 appears to be a sequence specific to the genus Drosophila and because of its different pharmacological properties to both Tyramine 1 and Tyramine 2 receptors, it is proposed that CG16766 be designated the founder member of a Tyramine 3 subgroup of receptors. Furthermore, the CG16766 sequence could potentially be used as a target site for the development of novel Drosophila-specific control agents (Bayliss, 2013).
It would thus appear that in insects tyramine could bring about its physiological actions by potentially interacting with two or three groups of 'tyramine receptors' and/or with the two additional groups of 'octopamine receptors', namely the α-adrenergic- and β-adrenergic-like octopamine receptors. To decide the exact physiological roles of tyramine in different locations in the insect nervous system, and in other tissues, it will be necessary to correlate the tissue-specific and stage-specific expression patterns of the above receptors with the release sites of both specific tyraminergic and octopaminergic neurons. Tyramine could potentially be released from both these types of neurones. Indeed, in a parallel situation in the CA1 field of the dorsal hippocampus, the release of dopamine from noradrenergic neurons has been suggested to be responsible for the activation of D1 dopamine receptors (Bayliss, 2013).
The Drosophila Genome Project database contains a gene, CG7431, annotated to be an "unclassifiable biogenic amine receptor." This gene has now been cloned and expressed in Chinese hamster ovary cells. After testing various ligands for G protein-coupled receptors, it was found that the receptor was specifically activated by tyramine (EC(50), 5x10-7M) and that it showed no cross-reactivity with β-phenylethylamine, octopamine, dopa, dopamine, adrenaline, noradrenaline, tryptamine, serotonin, histamine, and a library of 20 Drosophila neuropeptides (all tested in concentrations up to 10-5 or 10-4M). The receptor was also expressed in Xenopus oocytes, where it was, again, specifically activated by tyramine with an EC(50) of 3x10-7M. Northern blots showed that the receptor is already expressed in 8-hour-old embryos and that it continues to be expressed in all subsequent developmental stages. Adult flies express the receptor both in the head and body (thorax/abdomen) parts. In addition to the Drosophila tyramine receptor gene, CG7431, another closely related Drosophila gene, CG16766, was found that probably also codes for a tyramine receptor. Furthermore, similar tyramine-like receptor genes were found in the genomic databases from the malaria mosquito Anopheles gambiae and the honeybee Apis mellifera. These four tyramine or tyramine-like receptors constitute a new receptor family that is phylogenetically distinct from the previously identified insect octopamine/tyramine receptors. The Drosophila tyramine receptor is the first cloned insect G protein-coupled receptor that appears to be fully specific for tyramine (Cazzamali, 2005).
Search PubMed for articles about Drosophila TyrR and TyrII
Bayliss, A., Roselli, G. and Evans, P. D. (2013). A comparison of the signalling properties of two tyramine receptors from Drosophila. J Neurochem 125(1): 37-48.. PubMed ID: 23356740
Cazzamali, G., Klaerke, D. A. and Grimmelikhuijzen, C. J. (2005). A new family of insect tyramine receptors. Biochem Biophys Res Commun 338: 1189-1196. PubMed ID: 16274665
Chintapalli, V. R., Wang, J. and Dow, J. A. (2007). Using FlyAtlas to identify better Drosophila melanogaster models of human disease. Nat Genet 39(6): 715-720. PubMed ID: 17534367
El-Kholy, S., Stephano, F., Li, Y., Bhandari, A., Fink, C. and Roeder, T. (2015). Expression analysis of octopamine and tyramine receptors in Drosophila. Cell Tissue Res 361(3): 669-684.. PubMed ID: 25743690
Huang, J., Ohta, H., Inoue, N., Takao, H., Kita, T., Ozoe, F. and Ozoe, Y. (2009). Molecular cloning and pharmacological characterization of a Bombyx mori tyramine receptor selectively coupled to intracellular calcium mobilization. Insect Biochem Mol Biol 39(11): 842-849.. PubMed ID: 19833207
Huang, J., Liu, W., Qi, Y.X., Luo, J. and Montell, C. (2016). Neuromodulation of courtship drive through tyramine-responsive neurons in the Drosophila brain. Curr Biol 26(17):2246-56. PubMed ID: 27498566
Kutsukake, M., Komatsu, A., Yamamoto, D. and Ishiwa-Chigusa, S. (2000). A tyramine receptor gene mutation causes a defective olfactory behavior in Drosophila melanogaster. Gene 245(1): 31-42. PubMed ID: 10713442
Nagaya, Y., Kutsukake, M., Chigusa, S. I. and Komatsu, A. (2002). A trace amine, tyramine, functions as a neuromodulator in Drosophila melanogaster. Neurosci Lett 329: 324-328. PubMed ID: 12183041
Ormerod, K. G., Hadden, J. K., Deady, L. D., Mercier, A. J. and Krans, J. L. (2013). Action of octopamine and tyramine on muscles of Drosophila melanogaster larvae. J Neurophysiol 110: 1984-1996. PubMed ID: 23904495
date revised: 3 December 2016
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