No abstract

Alcohol intoxication at early ages is a risk factor for the development of addictive behavior. To uncover neuronal molecular correlates of acute ethanol intoxication, we used stable-isotope–labeled mice combined with quantitative mass spectrometry to screen more than 2,000 hippocampal proteins, of which 72 changed synaptic abundance up to twofold after ethanol exposure. Among those were mitochondrial proteins and proteins important for neuronal morphology, including MAP6 and ankyrin-G. Based on these candidate proteins, we found acute and lasting molecular, cellular, and behavioral changes following a single intoxication in alcohol-naïve mice. Immunofluorescence analysis revealed a shortening of axon initial segments. Longitudinal two-photon in vivo imaging showed increased synaptic dynamics and mitochondrial trafficking in axons. Knockdown of mitochondrial trafficking in dopaminergic neurons abolished conditioned alcohol preference in Drosophila flies. This study introduces mitochondrial trafficking as a process implicated in reward learning and highlights the potential of high-resolution proteomics to identify cellular mechanisms relevant for addictive behavior.

SummaryAlcohol intoxication at early ages is a risk factor for development of addictive behavior. To uncover neuronal molecular correlates of acute ethanol intoxication, we used stable-isotope labeled mice combined with quantitative mass spectrometry to screen over 2000 hippocampal proteins of which 72 changed synaptic abundance up to two-fold after ethanol exposure. Among those were mitochondrial proteins and proteins important for neuronal morphology, including MAP6 and Ankyrin-G. Based on these candidate proteins, we found acute and lasting molecular, cellular, and behavioral changes following a single intoxication in alcohol-naïve mice. Immunofluorescence analysis revealed a shortening of axon initial segments. Longitudinal two-photon in vivo imaging showed increased synaptic dynamics and mitochondrial trafficking in axons. Knockdown of mitochondrial trafficking in dopaminergic neurons abolished conditioned alcohol preference in Drosophila. This introduces mitochondrial trafficking as a process implicated in reward learning, and highlights the potential of high-resolution proteomics to identify cellular mechanisms relevant for addictive behavior.

A network of communicating tumour cells established by tumour microtubes (TMs) is supposed to mediate relevant aspects of progression and resistance of incurable gliomas. Moreover, neuronal activity has been shown to foster malignant behavior of glioma cells by non-synaptic paracrine and autocrine mechanisms. Here, we report an unexpected direct communication channel between neurons and glioma cells in multiple disease models as well as in astrocytomas and glioblastomas (GBs) of adult patients: functional bona fide chemical synapses formed between presynaptic neurons and postsynaptic glioma cells. These neurogliomal synapses (NGS) show a typical synaptic ultrastructure, are located on TM networks, and produce depolarizing postsynaptic currents mediated by glutamate receptors of the AMPA subtype. AMPA-type glutamate receptors (AMPAR) are expressed by a molecularly and morphologically distinct subpopulation of network-integrated glioma cells. Increased neuronal activity under epileptic conditions ex vivo or neuronal optogenetic stimulation in vivo enhanced, while general anesthesia diminished synchronized calcium transients in TM-connected glioma networks. Accordingly, anesthesia reduced invasiveness of TM-positive tumour cells in mice. Genetic perturbation of AMPAR or chronic AMPAR inhibition by perampanel decreased glioma invasion and proliferation in mice and deletion of GluRII in Drosophila glioma increased survival. These findings reveal a hitherto unappreciated direct synaptic communication between neurons and glioma cells that appears relevant for brain tumour biology, implying new avenues for glioma treatment.

Glioblastoma are incurable brain tumors characterized by their colonization of the entire brain and their notorious therapeutic resistance. Recently, we discovered long membrane tubes called tumor microtubes contributing to invasion, network formation of tumor-tumor networks and therapeutic resistance. Subsequently, heterogeneous networks of neurons and glioblastoma cells were characterized, which can communicate by synaptic and perisynaptic contacts as well as by paracrine mechanisms. Currently used models of studying neuron-glioblastoma interactions are limited by the possibility to study glioblastoma in a defined human neuronal microenvironment. Here, we set out to derive excitatory and inhibitory neurons from embryonic stem cells via lentiviral reprogramming and co-cultured them with patient-derived glioblastoma cells. We could show that structural and functional neuron-glioblastoma synaptic contacts are formed. Functional communication between neurons and glioblastoma cells were characterized with calcium imaging, showing similar complex calcium dynamics previously characterized with in vivo imaging of patient-derived xenograft models. The single-cell glioblastoma morphology was morphometrically similar to that of human glioblastoma tissue. Tumor microtubes and the formation of tumor-tumor networks could be demonstrated. Additionally, glioblastoma invasion patterns in our human neuronal co-culture model resemble invasion patterns recently characterized with patient-derived xenograft models. Lastly, we investigated reciprocal neuron-glioblastoma interactions and longitudinally characterized neuronal activity with patch-clamp electrophysiology. In conclusion, we provide a novel human neuron-glioblastoma co-culture system allowing in-depth molecular and functional characterization for future Cancer Neuroscience studies.