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Si us plau utilitzeu sempre aquest identificador per citar o enllaçar aquest document: https://hdl.handle.net/2445/223879
Novel pharmacological strategies for neurological conditions involving glutamate and adenosine
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[eng] Neuropsychiatric disorders such as schizophrenia and bipolar disorder present alterations in dopaminergic and glutamatergic neurotransmission, which has led to the development of therapeutic strategies targeting these systems. Recently, the adenosynergic system has emerged as a key regulator of dopaminergic and glutamatergic activity, with evidence that activation of adenosine receptorA2A can modulate dopaminergic transmission and affect neuropsychiatric symptoms. In addition, the interaction between the A2A receptor and the metabotropic glutamate receptor type 5 in the striatum represents a potential therapeutic convergence point for the treatment of these disorders. In this context, nanobodies have emerged as innovative tools for modulating membrane macromolecular interactions in a highly specific manner. Unlike conventional antibodies, nanobodies are heavy-chain antibody fragments that have advantages such as their small size, high stability, and ability to access inaccessible regions of proteins. In this thesis, we explore the development and characterization of the Nb43 nanobody, designed to modulate the glutamate receptor mGlu5R, as well as its Nb43-CGS21680 conjugate, which incorporates an A2AR agonist to enhance its effect. The thesis is divided into three chapters. In the first chapter, we characterize in vitro the binding and functional effect of Nb43 and Nb43-CGS21680 in cellular systems expressing mGlu5R and A2AR. In the second chapter, we evaluate the distribution, penetration into the central nervous system and pharmacological activity of these nanobodies in murine models. Finally, in the third chapter, we performed a meta-analysis of adenosine metabolism modulators in schizophrenia and bipolar disorder, to evaluate their clinical impact and therapeutic potential. The results of the in vitro study of Nb43 showed that Nb43 selectively binds to mGlu5R in a physiological context. On the other hand, Nb43-CGS21680 showed a dual-action profile, with simultaneous modulation of mGlu5R and A2AR. Interestingly, the co-activation of these receptors resulted in a more specific modulation of intracellular activity. In vitro experiments of PLA and NanoBiT confirmed the existence of functional interactions between mGlu5R and A2AR, supporting the hypothesis that these receptors form heteromers in the neuronal membrane. To evaluate the biodistribution of Nb43 and Nb43-CGS21680, we performed in vivo imaging studies after intravenous and intranasal administration. The results indicated that Nb43 does not cross the blood-brain barrier significantly when administered intravenously. However, in intranasal administration, small concentrations were detected in the olfactory bulb, suggesting a possible route of access to the central nervous system. The behavioral effects of Nb43 and Nb43-CGS21680 were evaluated after their intracerebroventricular administration, which allowed analyzing their direct impact on the central nervous system without the limitation of the blood-brain barrier. In the locomotion test, locomotor activity CGS21680 significantly reduced, in line with previous studies suggesting its anti-dopaminergic effect. Similarly, Nb43 and Nb43-CGS21680 also decreased locomotion, although with less intensity. This effect suggests that Nb43 could be modulating dopaminergic neurotransmission through its action on mGlu5R in the striatum. In the haloperidol-induced catalepsy test, catalepsy CGS21680 significantly increased, confirming its role in dopaminergic inhibition. Interestingly, Nb43 also exacerbated catalepsy, indicating that its effect on mGlu5R could be modulating the activity of the dopaminergic pathway. Unlike CGS21680, whose effects disappeared after 24-48 hours, Nb43-CGS21680 prolonged the catalepsy effect until 48 hours after administration, suggesting a more sustained mechanism of modulation of neurotransmission. Additionally, we performed catalepsy experiments after intranasal administration of Nb43 and Nb43-CGS21680. The results showed that Nb43 had a significant impact on haloperidol-induced catalepsy, while Nb43-CGS21680 showed no effects. This suggests that Nb43 was able to reach the central nervous system through the olfactory bulb, where mGlu5R is expressed and its role in modulating locomotion and balance has been reported. On the other hand, the meta-analysis study included clinical studies that evaluated the impact of adenosine metabolism modulators, such as allopurinol, dipyridamole, pentoxifylline, and propentophylline, on the symptomatology of schizophrenia and bipolar disorder. The results indicated that allopurinol, a xanthine oxidase inhibitor that raises adenosine levels, was the most studied and showed positive effects in reducing positive and negative symptoms of schizophrenia and bipolar disorder. Dipyridamole, an adenosine reuptake inhibitor, also showed improvements in positive symptoms, although studies were limited. Propentophylline, a xanthine derivative with neuroprotective and anti-inflammatory, had more marked effects on negative symptoms and cognitive impairment. The findings of the meta-analysis reinforce the adenosynergistic hypothesis of schizophrenia and bipolar disorder, which proposes that modulation of the adenosynergistic system may influence dopaminergic and glutamatergic neurotransmission. In this sense, the results suggest that strategies aimed at enhancing adenosynergistic signaling could represent a promising therapeutic avenue. Overall, this thesis contributes to the knowledge about the role of nanobodies in the pharmacology of membrane receptors and highlights the importance of the adenosynergic system in the modulation of dopaminergic and glutamatergic neurotransmission in neuropsychiatric disorders.
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SARASOLA TELLERIA, Laura i.. Novel pharmacological strategies for neurological conditions involving glutamate and adenosine. [consulta: 6 de desembre de 2025]. [Disponible a: https://hdl.handle.net/2445/223879]