magistrsko delo
Nika Kozoderc (Avtor), Marko Kreft (Recenzent), Nina Vardjan (Mentor)

Povzetek

Nevroni in celice glije na svojih membranah izražajo številne z G-proteini sklopljene receptorje (GPCR), ki imajo pomembno vlogo pri uravnavanju celičnih procesov v možganih. Noradrenalin (NA) je nevromodulator v osrednjem živčnem sistemu (OŽS) vretenčarjev, ki ga izločajo noradrenergični nevroni s telesi v možganskem jedru locus coeruleus (LC). NA se veže na adrenergične receptorje (AR) možganskih celic, ki so GPCR, kar odvisno od podtipa AR in sklopitve z Gα-podenoto (Gαq, Gαs, Gαi, Gα12/13) G-proteina, vodi v aktivacijo Ca2+- in/ali cAMP-signalnih poti. LC-noradrenergični sistem ključno sodeluje pri uravnavanju presnove, pozornosti, učenja in tvorbe spomina ter drugih fizioloških procesov v OŽS. Delovanje le-tega je s staranjem in pri nevrodegeneraciji okrnjeno, kar lahko vpliva na zmanjšanje kognitivnih sposobnosti. V OŽS vinske mušice analogno vlogo NA opravljata nevromodulatorja oktopamin (OA) in njegov prekurzor tiramin (TA). Obstajajo številne strukturne, signalizacijske in funkcijske podobnosti med NA v vretenčarjih in OA in TA v nevretenčarjih, prav tako med AR in receptorji za OA in/ali TA (OAR, TAR). Namen naloge je bil ugotoviti, kakšen vpliv ima TA in/ali OA na raven znotrajcelične koncentracije Ca2+ in cAMP v nevronih in korteksni gliji v možganih vinske mušice. Z uporabo genetsko kodirajočega Ca2+-indikatorja jGCaMP7b smo spremljali spremembo znotrajcelične koncentracije Ca2+ in s FRET-nanosenzorjem Epac1-camps spremembo znotrajcelične koncentracije cAMP v nevronih oz. korteksni gliji v možganih vinske mušice s konfokalnim mikroskopom v realnem času. Ugotovili smo, da mešanica OA in TA povzroči porast v znotrajcelični koncentraciji Ca2+ v nevronih in korteksni gliji najverjetneje preko aktivacije z Gαq-proteini-sklopljenih OAR in TAR na površini celic. Porast v Ca2+ je bil v nevronih 9,8-krat višji kot v korteksni gliji, kar nakazuje na večjo Ca2+-vzdražnost nevronov v možganih vinske mušice po oktopaminergični/tiraminergični aktivaciji. Mešanica OA in TA je povzročila porast v znotrajcelični koncentraciji cAMP samo v nevronih, medtem ko v korteksni gliji porasta nismo opazili. Porast v cAMP v nevronih je najverjetneje posledica aktivacije z Gαs-proteini-sklopljenih OAR in TAR na površini nevronov. Draženje možganov mladih vinskih mušic z OA je povzročilo porast v Ca2+ v korteksni gliji, česar v možganih staranih vinskih mušic nismo opazili, kar nakazuje na okvaro z OA-posredovane Ca2+-signalizacije. Pri draženju možganov mladih kot staranih vinskih mušic z OA nismo zaznali sprememb v cAMP-odzivu. Okvara Ca2+-signalizacije v celicah glije, ki v OŽS skrbijo za homeostazo, bi lahko v možganih starajočih živali prispevala h kognitivnemu upadu živali in bi v prihodnje lahko predstavljala novo tarčo za razvoj zdravil pri nevrodegenerativnih obolenjih.

Ključne besede

vinska mušica;Ca2+;cAMP;nevroni;korteksna glija;oktopamin;tiramin;konfokalna mikroskopija;staranje;

Podatki

Jezik: Slovenski jezik
Leto izida:
Tipologija: 2.09 - Magistrsko delo
Organizacija: UL BF - Biotehniška fakulteta
Založnik: [N. Kozoderc]
UDK: 595.772.2:572.788(043.2)
COBISS: 189190659 Povezava se bo odprla v novem oknu
Št. ogledov: 33
Št. prenosov: 5
Ocena: 0 (0 glasov)
Metapodatki: JSON JSON-RDF JSON-LD TURTLE N-TRIPLES XML RDFA MICRODATA DC-XML DC-RDF RDF

Ostali podatki

Sekundarni jezik: Angleški jezik
Sekundarni naslov: Measurements of intracellular concentration of Ca[sup]2+ and cAMP in brain cells of fruit fly upon octopamine and tyramine stimulation
Sekundarni povzetek: Neurons and glial cells express numerous G-protein-coupled receptors (GPCRs) on their membranes, which play an important role in regulating cellular processes in the brain. Noradrenaline (NA), a neuromodulator in the vertebrate central nervous system (CNS), is secreted by noradrenergic neurons, which bodies originate in the nucleus locus coeruleus (LC). NA binds to adrenergic receptors (AR) on brain cells, which are GPCRs, and depending on the AR subtype and coupling with the Gα-subunit (Gαs, Gαq, Gαi, Gα12/13) of G-protein, the latter leads to the activation of Ca2+- and/or cAMP-signaling pathways. The LC-noradrenergic system plays a key role in the regulation of metabolism, attention, learning and memory formation, as well as other physiological processes in the CNS. Its functioning is impaired with aging and neurodegeneration, which can project in cognitive decline. Octopamine (OA) and its precursor tyramine (TA), neuromodulators in the CNS of the fruit fly, have analogous functions to those of NA in vertebrates. There are many structural, signaling and functional similarities between NA in vertebrates and OA and TA in invertebrates as well as between AR and OA and/or TA receptors (OAR, TAR). The purpose of the task was to determine the effect of TA and/or OA on the intracellular Ca2+ and cAMP concentration in neurons and cortical glia in the fruit fly brain. With the genetically encoded Ca2+-indicator jGCaMP7b and FRET-nanosensor Epac1-camps we monitored the change in the intracellular concentration of Ca2+ and cAMP in neurons or cortical glia in the fruit fly brain in real-time with confocal microscopy. We found that the mixture of OA and TA causes an increase in the intracellular Ca2+ concentration in neurons and cortical glia, most likely through the activation of Gαq-protein-coupled OARs and TARs. The increase in Ca2+ was 9,8-fold higher in neurons than in cortical glia, suggesting a higher Ca2+ excitability of neurons in the fruit fly brain after octopaminergic/tyraminergic activation. The mixture of OA and TA caused an increase in intracellular cAMP levels only in neurons, while no increase was observed in cortical glia. The increase in cAMP in neurons is most likely due to the activation of Gαs-protein-coupled OARs and TARs. Stimulation of the brain of young fruit flies with OA caused an increase in Ca2+ in cortical glia, which was not observed in the brain of aged fruit flies, suggesting an impairment of OA-mediated Ca2+-signaling. Upon stimulation of the brain of aged fruit flies with OA, no changes in the cAMP response were observed in cortical glia, as in young flies. Impairment of Ca2+-signaling in glial cells, which are responsible for maintaining CNS homeostasis, may contribute to cognitive decline and represent a new target for the development of novel drugs against neurodegenerative diseases in the future.
Sekundarne ključne besede: fruit fly;Ca2+;cAMP;neurons;cortical glia;octopamine;tyramine;confocal microscopy;aging;
Vrsta dela (COBISS): Magistrsko delo/naloga
Študijski program: 0
Konec prepovedi (OpenAIRE): 1970-01-01
Komentar na gradivo: Univ. v Ljubljani, Biotehniška fak., Oddelek za biologijo, Magistrsko delo magistrskega študijskega programa 2. stopnje Molekulske in funkcionalne biologije
Strani: 1 spletni vir (1 datoteka PDF (XIII, 77 str.)
ID: 23140489