Astrocyte modulation of synaptic transmission in the reward circuitry of the Ventral Tegmental Area

Abstract (inglese) The mesocorticolimbic dopaminergic (DA) system originating from the Ventral Tegmental Area (VTA) plays a prominent role in the cognitive processing of aversion, motivation, pleasure and reward, including the development of addiction. A central aspect for the function of the brain reward system is the switch in the pattern of action potential discharges of VTA DA neurons from a tonic, low firing frequency to a phasic, high frequency bursts which cause both an increased DA release in the VTA projecting structures and a local release of endocannabinoids (eCBs). Glutamatergic signalling to the VTA plays a central role in this transition from tonic to phasic-burst firing pattern and long-term changes of this signaling pathway can profoundly alter the output of DA neurons. Over the last years, astrocytes emerged as important regulatory elements of synaptic transmission in different brain circuits. They respond to neurotransmitters with Ca2+ elevations through a mechanism that involves the production of inositol-1,4,5-trisphosphate (IP3) and the release of Ca2+ from IP3-sensitive intracellular Ca2+ stores. In turn, these Ca2+ elevations in astrocytes evoke the release of gliotransmitters that modulates synaptic transmission. Whether a similar mechanism operates in VTA networks remains unexplored. In my thesis, by combining patch-clamp recording techniques and Ca2+ imaging experiments in horizontal VTA slices of both juvenile (P14-17) and young adult (P30-70) mice, we investigated whether astrocytes contribute to the modulation of excitatory synaptic transmission in the reward VTA circuitry. The results described in the present thesis demonstrate that astrocyte signaling contributes to long-term plastic changes of glutamatergic transmission in VTA circuitry. Our study opens a new perspective for the understanding of the cellular and molecular mechanisms that control the brain reward system.  Abstract (italiano) L’ Area Tegmentale ventrale (VTA) è una piccola regione cerebrale, da cui ha origine il sistema mesolimbico-corticale che gioca un ruolo fondamentale in importanti processi cognitivi quali il piacere, la motivazione e la ricompensa. I neuroni dopaminergici, che rappresentano la popolazione cellulare maggiormente rappresentata in VTA, possiedono peculiari caratteristiche elettrofisiologiche. Essi, infatti possiedono un’ attività tonica a livello basale che puo' improvvisamente tradursi in una di tipo fasico, o a burst, al presentarsi di situazioni comportamentali salienti, quali ad esempio, l’ottenimento di una ricompensa inaspettata e gratificante. Una delle conseguenze di tale cambio di attività nei neuroni dopaminergici è un maggiore rilascio di dopamina a livello delle strutture bersaglio e il rilascio di endocannabinoidi a livello locale. Gli inputs glutammatergici alla VTA giocano un ruolo fondamentale nel cambio di attività di questi neuroni. La plasticità della trasmissione glutammatergica è dunque importante nel determinare l’output dei neuroni dopaminergici stessi. Negli ultimi anni gli astrociti si sono rivelati essere importanti elementi regolatori in diverse funzioni cerebrali. Essi infatti rispondono ai neurotrasmettitori rilasciati dai neuroni con aumenti intracellulari dello ione Ca2+, attraverso meccanismi che coinvolgono la produzione di inositolo-1,4,5-trisphosphate (IP3) e rilascio di Ca2+ dagli stores intracellulari. Gli aumenti del Ca2+ degli astrociti regolano, a loro volta, il rilascio di gliotrasmettitori che possono modulare la trasmissione sinaptica. Tuttavia, non è noto se tali meccanismi siano operativi anche nella VTA. L’obbiettivo della mia tesi è quello di studiare una possibile modulazione della trasmissione sinaptica nella VTA da parte degli astrociti. Per caratterizzare questa interazione ho accoppiato registrazioni elettrofisiologiche a studi di imaging del Ca2+ in fettine di VTA ottenute da animali giovani (P14-17) e da animali adulti (P30-70). I risultati ottenuti nella presente tesi suggeriscono che gli astrociti contribuiscono alla modifica a lungo termine della trasmissione sinaptica glutammatergica nel circuito della VTA. Tali risultati aprono nuove prospettive nella comprensione dei meccanismi modulatori presenti nei circuiti della ricompensa.

Dynamic interactions between GABAergic and astrocytic networks

Brain network activity derives from the concerted action of different cell populations. Together with interneurons, astrocytes play fundamental roles in shaping the inhibition in brain circuitries and modulating neuronal transmission. In this review, we summarize past and recent findings that reveal in neural networks the importance of the interaction between GABAergic signaling and astrocytes and discuss its physiological and pathological relevance

GABA tonic currents and glial cells are altered during epileptogenesis in a mouse model of Dravet syndrome

Dravet Syndrome (DS) is a rare autosomic encephalopathy with epilepsy linked to Na(v)1.1 channel mutations and defective GABAergic signaling. Effective therapies for this syndrome are lacking, urging a better comprehension of the mechanisms involved. In a recognized mouse model of DS, we studied GABA tonic current, a form of inhibition largely neglected in DS, in brain slices from developing mice before spontaneous seizures are reported. In neurons from the temporal cortex (TeCx) and CA1 region, GABA tonic current was reduced in DS mice compared to controls, while in the entorhinal cortex (ECx) it was not affected. In this region however allopregnanonole potentiation of GABA tonic current was reduced in DS mice, suggesting altered extrasynaptic GABA(A) subunits. Using THIP as a selective agonist, we found reduced δ subunit mediated tonic currents in ECx of DS mice. Unexpectedly in the dentate gyrus (DG), a region with high δ subunit expression, THIP-evoked currents in DS mice were larger than in controls. An immunofluorescence study confirmed that δ subunit expression was reduced in ECx and increased in DG of DS mice. Finally, considering the importance of neuroinflammation in epilepsy and neurodevelopmental disorders, we evaluated classical markers of glia activation. Our results show that DS mice have increased Iba1 reactivity and GFAP expression in both ECx and DG, compared to controls. Altogether we report that before spontaneous seizures, DS mice develop significant alterations of GABA tonic currents and glial cell activation. Understanding all the mechanisms involved in these alterations during disease maturation and progression may unveil new therapeutic targets

Interneuron-specific signaling evokes distinctive somatostatin-mediated responses in adult cortical astrocytes.

The signaling diversity of GABAergic interneurons to post-synaptic neurons is crucial to generate the functional heterogeneity that characterizes brain circuits. Whether this diversity applies to other brain cells, such as the glial cells astrocytes, remains unexplored. Using optogenetics and two-photon functional imaging in the adult mouse neocortex, we here reveal that parvalbumin- and somatostatin-expressing interneurons, two key interneuron classes in the brain, differentially signal to astrocytes inducing weak and robust GABAB receptor-mediated Ca2+ elevations, respectively. Furthermore, the astrocyte response depresses upon parvalbumin interneuron repetitive stimulations and potentiates upon somatostatin interneuron repetitive stimulations, revealing a distinguished astrocyte plasticity. Remarkably, the potentiated response crucially depends on the neuropeptide somatostatin, released by somatostatin interneurons, which activates somatostatin receptors at astrocytic processes. Our study unveils, in the living brain, a hitherto unidentified signaling specificity between interneuron subtypes and astrocytes opening a new perspective into the role of astrocytes as non-neuronal components of inhibitory circuits

Interneuron-specific signaling evokes distinctive somatostatin-mediated responses in adult cortical astrocytes

The signaling diversity of GABAergic interneurons to post-synaptic neurons is crucial to generate the functional heterogeneity that characterizes brain circuits. Whether this diversity applies to other brain cells, such as the glial cells astrocytes, remains unexplored. Using optogenetics and two-photon functional imaging in the adult mouse neocortex, we here reveal that parvalbumin- and somatostatin-expressing interneurons, two key interneuron classes in the brain, differentially signal to astrocytes inducing weak and robust GABAB receptor-mediated Ca2+ elevations, respectively. Furthermore, the astrocyte response depresses upon parvalbumin interneuron repetitive stimulations and potentiates upon somatostatin interneuron repetitive stimulations, revealing a distinguished astrocyte plasticity. Remarkably, the potentiated response crucially depends on the neuropeptide somatostatin, released by somatostatin interneurons, which activates somatostatin receptors at astrocytic processes. Our study unveils, in the living brain, a hitherto unidentified signaling specificity between interneuron subtypes and astrocytes opening a new perspective into the role of astrocytes as non-neuronal components of inhibitory circuits

Astrocytes mediate long-lasting synaptic regulation of ventral tegmental area dopamine neurons

This study shows that burst-firing-induced astrocyte signaling potentiates glutamatergic transmission onto dopamine neurons in mouse VTA.The plasticity of glutamatergic transmission in the ventral tegmental area (VTA) represents a fundamental mechanism in the modulation of dopamine neuron burst firing and phasic dopamine release at target regions. These processes encode basic behavioral responses, including locomotor activity, learning and motivated behaviors. Here we describe a hitherto unidentified mechanism of long-term synaptic plasticity in mouse VTA. We found that the burst firing in individual dopamine neurons induces a long-lasting potentiation of excitatory synapses on adjacent dopamine neurons that crucially depends on Ca2+ elevations in astrocytes, mediated by endocannabinoid CB1 and dopamine D2 receptors co-localized at the same astrocytic process, and activation of pre-synaptic metabotropic glutamate receptors. Consistent with these findings, selective in vivo activation of astrocytes increases the burst firing of dopamine neurons in the VTA and induces locomotor hyperactivity. Astrocytes play, therefore, a key role in the modulation of VTA dopamine neuron functional activity