20.500.12556/RUL-150013 Vzdražnost celic v možganih vinske mušice Drosophile melanogaster po oktopaminergični aktivaciji Excitability of cells in the brain of the fruit fly Drosophila melanogaster upon octopaminergic activation Nevroni so električno vzdražne celice, medtem ko so astrociti, podskupina glija celic, električno nevzdražne celice. Astrociti zaznavajo aktivnosti v možganih z receptorji, ki so na plazemski membrani. Z G-proteini sklopljeni receptorji (GPCR) so metabotropni receptorji, ki so izraženi tako na astrocitih kot na nevronih. Podskupina GPCR, značilna za vretenčarje, so adrenergični receptorji, tipa α (α1, α2) in ß (ß1-3), ki se odzivajo v možganih na stresni nevromodulator noradrenalin. V vinski mušici Drosophili melanogaster, in drugih nevretenčarjih, vlogo (nor)adrenergičnega sistema opravlja oktopaminergični/tiraminergični sistem. Signalni molekuli oktopaminergičnega/tiraminergičnega sistema sta monoamina oktopamin in tiramin, ki delujeta preko aktivacije oktopaminskih in/ali oktopamin/tiraminskih receptorjev, kar sproži aktivacijo Ca2+– in cAMP–signalnih poti. Ker vpliv oktopamina na znotrajcelično raven Ca2+ v možganskih celicah vinske mušice še ni popolnoma raziskan, smo želeli ugotoviti, ali se nevroni v možganih vinske mušice odzovejo na draženje z oktopaminom s porastom v znotrajcelični koncentraciji Ca2+, ki je koncentracijsko odvisen, in ali je Ca2+-vzdražnost nevronov v možganih vinske mušice po draženju z oktopaminom drugačna kot v nevrogliji, kot so astrociti. Z uporabo genetsko kodirajočega fluorescenčnega senzorja za Ca2+, jGCaMP7b, in konfokalno mikroskopijo v realnem času smo ugotovili, da se nevroni odzovejo na draženje z oktopaminom s porastom v znotrajcelični koncentraciji Ca2+, ki je koncentracijsko odvisen. Po izračunu polovične efektivne koncentracije (EC50) oktopamina za nevrone, ki je znašala 10 μM, smo dražili astrocite z 10 μM in rezultate primerjali z nevroni. Astrociti so se odzvali na 10 μM oktopamin s porastom znotrajcelične koncentracije Ca2+, vendar je bil porast 9-krat nižji kot v nevronih. Po drugi strani pa je bila EC50 za oktopamin v nevronih 10-krat višja kot v astrocitih, kar nakazuje, da je Ca2+-vzdražnost na oktopaminergični dražljaj v nevronih nižja kot v astrocitih. Neurons are electrically excitable cells, while astrocytes, a subset of neuroglia cells, are electrically non-excitable cells. Astrocytes detect activities in the brain with receptors on the plasma membrane. G-protein coupled receptors (GPCRs) are metabotropic receptors that are expressed on both astrocytes and neurons. A subgroup of GPCRs specific to vertebrates are adrenergic receptors, type α (α1, α2) and ß (ß1-3), which respond in the brain to the stress neuromodulator noradrenaline. In the fruit fly Drosophila melanogaster and other invertebrates, the role of the (nor)adrenergic system is performed by the octopaminergic/tyraminergic system. The signaling molecules of the octopaminergic/tyraminergic system are the monoamines octopamine and tyramine, which act through the activation of octopamine and/or octopamine/tyramine receptors, which triggers the activation of Ca2+ and cAMP signaling pathways. As the effect of octopamine on the intracellular Ca2+ levels in the fruit fly brain cells is not yet fully understood, we wanted to determine whether neurons respond to octopamine stimulation with a concentration-dependent increase in intracellular Ca2+ concentration and whether the Ca2+-excitability of neurons in the fruit fly brain upon octopamine stimulation is different from that of neuroglia, such as astrocytes. Using a genetically encoded fluorescent Ca2+ sensor, jGCaMP7b, and real-time confocal microscopy, we found that neurons respond to octopamine stimulation with a concentration-dependent increase in intracellular Ca2+ concentration. After calculating the half-effective concentration (EC50) of octopamine for neurons, which was 10 μM, we stimulated astrocytes with 10 μM octopamin and compared the Ca2+-response with neurons. Astrocytes responded to 10 μM octopamine with an 9-fold lower increase in intracellular Ca2+ concentration than that in neurons. In contrast, the EC50 for octopamine was 10-fold higher in neurons than astrocytes, suggesting that Ca2+-excitability to the octopaminergic stimulus is lower in neurons than astrocytes. Drosophila melanogaster nevroni astrociti oktopamin Ca2+ EC50 konfokalna mikroskopija Drosophila melanogaster neurons astrocytes Ca2+ octopamine EC50 confocal microscopy true false false [E. Sanjković] Slovenski jezik Angleški jezik Magistrsko delo/naloga 2023-09-13 08:15:29 2023-09-13 08:15:32 2023-09-15 03:18:54 0000-00-00 00:00:00 2023 0 0 0000-00-00 NiDoloceno NiDoloceno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 1970-01-01 575(043.2) 229173 164359171 34812.pdf 34812.pdf 1 6B8D39C01EE029C4418DAEA9BD04E5F1 e067772eddea81fe4255626fc9d8dc8ba193571b2ce095fd2ced43278cbf1f60 f386cd38-51fc-11ee-916e-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=174546 Biotehniška fakulteta 0 0 0