The cells of multicellular organisms developed various ways of communication for maintaining homeostasis and response to their inner and outer environment [1]. In addition to endocrine, paracrine, juxtacrine and autocrine signaling, the cells send signals via extracellular membrane bound structures, i.e. extracellular vesicles, and can be classified depending on their size, molecular composition and synthesis pathways [2]. The cells within central nervous system uses these extracellular vesicles for long distance communication, which allows nucleic acid (both DNA and different types RNA), protein and special lipid domain transfer between the cells [3].

The extracellular vesicles can be classified as exosomes, which has a diameter ranging from 30 nm to 100 nm; microvesicles with diameter between 100-1000 nm; apoptotic bodies, which are released from dying cells via apoptotic process, and has a diameter between 800-5000 nm, and large oncosomes, which are released from neoplastic cells and has a diameter larger than 1 μm and can be large as 10 μm [4,5].

The exosomes are generated via multivesicular bodies (MVB) which are derived from early endosomes. Early endosomes contain lipid rafts with specific proteins e.g. tetraspanins, adhesion, fusion, and receptor transport proteins. With the intraluminal vesicle (ILV) formation, these molecules are incorporated into the exosomal membrane [6]. ESCRT, TSG-101, Alix proteins are responsible for the sorting of exosomes and after packaging, exosomes expelled to the extracellular domain via a fusion process led by Rab GTPases (Figure 1) [7].

Microvesicles are the generated through the budding of plasma membrane, which is regulated through translocation of phospholipids by floppase/flippase enzymes, organization of cytoskeleton under plasma membrane with ADP-ribosylation factor 6 (ARF6), Phospholipase D (PLD), Extracellularsignal regulated kinase (ERK) and Myosin light-chain kinase (MLCK) enzymes [8]. The molecules participating into extracellular vesicle biogenesis process are candidates for EV markers, which useful in detection and isolation of EVs (Figure 2) [9].

EVs can be isolated from serum, saliva, cerebrospinal fluid (CSF), urine, breast milk, synovial fluid, effusions, semen and cell cultures [10]. The isolation of EVs requires either different centrifuge steps up to an ultracentrifuge with 100,000 g and/or mechanical filters, which are specifically designed according to diameter preferred EV subtype [11]. The EV purification and subpopulation selection can be performed with immunocapture methods [12]. The isolated EVs can be imaged with transmission and/or scanning electron microscopes. Transmission electron microscopy can be combined with immunogold method to show a specific protein within or on the EV [13]. The light microscopes with capability of super-resolution can be recruited for the imaging of EVs, particularly for exosomes, which has a diameter smaller than 100 nm [14]. The isolated EVs can be analyzed for their content of proteins, nucleic acids, lipid and carbohydrates [15].

Since CNS is sealed by bones, connective tissues and bloodbrain barriers, also contains vital centers for cognition, sensory, motor and autonomic functions, the indications for taking direct biopsies for diagnostic purposes is limited only to some life threating conditions like infection/abscess or brain tumors [16]. Many central nervous system pathologies are diagnosed with imaging modalities, including computed tomography (CT) and magnetic resonance imaging (MRI) [17]. Through these imaging modalities are capable of noninvasively monitoring the organs within CNS, generally they don’t any functional information regarding cellular processes [18]. The functional imaging modalities like functional MRI (fMRI) or positron emission tomography (PET) shows the metabolic activity in brain, however these techniques still suffers the resolution problem, since they can’t monitor individual neuronal activity [19]. The best technique for gaining biochemical, cellular or microbiological information from CNS is obtaining CSF via lumbar puncture [20]. EVs can be collected form CSF and provide valuable information regarding the functional status of neurons and glial cells within the CNS [21]. However, lumbar puncture is still an invasive method with its possible complications e.g. hemorrhage, spinal cord/nerve root injury, even cerebral hernia [22]. In addition, the content of CSF is modified through the components of CSF- brain barrier, i.e. choroid plexus/ ependymal cells, neural and glial processes [23]. Exosomes from CNS carry molecules originating from the inner compartment, including ILV and EBV machinery, of their source cells (neurons, glial cells). In addition, these vesicles carry the inside information and messages from the CNS cells, which reside in a sealed biological compartment. It can be hypothesized, that the information within the exosomes is changed minimally during their journey from CNS to the peripheral blood. Thus, exosomes might contain more proper information regarding cellular/metabolic status within CNS when they are compared to CSF.

An alternative and relatively noninvasive way to obtain information from CNS is to collect EVs which are shed from blood brain barrier [24]. EVs from blood brain barrier contains EVs from both CNS and blood brain barrier components, i.e. endothelial cells and astrocytes [25]. The EVs from neurons and glial cells can cross blood brain barrier via transcytosis process [26]. The main limitations for obtaining EVs from peripheral blood sample are isolation of blood brain barrier derived EVs from peripheral EVs and the determination of origins of EVs derived from blood brain barrier. The areas where sealing of blood brain barrier is relatively weak (e.g. circumventricular organs) might be comparatively more direct source for exosome originating from CNS [27]. In addition, EVs with neuronal origins can be isolated from peripheral blood samples through various surface markers like NCAM or L1CAM [28]. Exosomes with neural origins also carry GLP-anchored prion protein and Glutamate receptor GluR2/3 subunits. Since these proteins are shown in neural cell culture as neural exosomal markers, more research is needed for their use for peripheral blood neural exosome isolation [29,30]. The proteomic analysis of exosomes derived from microglia culture showed the expression of CD13 and lactate transporter MCT-1 [31]. The exosomes with oligodendroglia origin has myelin components, like myelin basic protein or myelin oligodendrocyte glycoprotein [32]. Another approach for the isolation of CNS derived exosomes is the use lipidomic and proteomic profiles of these exosomes. The lipid composition of exosomes (phosphatidylserine, sphingomyelin, lyso-phosphatidylcholine, lysophosphatidylethanolamine and other families), provide information regarding the origin of exosomes [33].

The isolation and biochemical analysis of EVs in psychiatric disorders can offer wide diagnostic and therapeutic opportunities, in addition to provision of valuable information regarding pathophysiology of these diseases [34]. In current clinical practice, imaging of CNS is not in routine use for the differential diagnosis of psychiatric diseases [35]. The blood tests are usually recruited for monitoring drug levels, ruling out endocrine diseases, micronutrient deficiencies or organ failures which might affect the mental status of patients [36,37]. However, blood tests are not capable to demonstrate the central nervous system biochemical and cellular properties. The information regarding the level of neurotransmitters, neural metabolic status, epigenetic and/or genetic alterations in neurons would provide valuable information for diagnosis and individualization of treatment of psychiatric disorders.

Nevertheless, the previous studies focusing on EVs in psychiatric disorders are predominantly executed in postmortem brain tissues of the patients [38-41]. The use of EVs extracted from peripheral blood samples for diagnostic purposes requires further basic research and clinical studies carried on patients diagnosed with psychiatric disorders. However, these preceding ex-vivo (postmortem) studies might guide the researchers to find potential targets to evaluate in patient population in order to develop biomarkers for diagnostic and/or therapeutic purposes.

In this part of the review, we will focus on some important psychiatric disorders, in which the use of EVs might be beneficial in terms of basic research, diagnosis (including secondary prevention of psychiatric disorders) and treatment. We decided to review EVs in depression, bipolar disorder, schizophrenia, and Alzheimer’s disease in the context of Research Domain Criteria (RDoC), since the constructs of these criteria attempt to reflect the etiology of psychiatric diseases in different dimension, extending from the molecules to the neural circuitries and anatomical structures [42]. Although the previous studies on EV in psychiatric disorders do not cover all aspects of RDoC criteria, understanding of EV characteristics in relation to a specific RDoC construct is still important to clarify the pathogenesis and finding appropriate biomarkers for these disorders.

The RDoC constructs provide valuable information for choosing candidates for biomarker molecules for both diagnostic and therapeutic purposes in psychiatric disorders. By using methods of bioinformatics, these potential biomarkers can be extracted from RDoC domain matrices. Extracellular vesicles provide a chance for measuring different classes of biomarkers (e.g. miRNA, proteins and lipids) simultaneously, which might increase the predictive values of testing and help to overcome the difficulties of working with a diagnostic modality relying on a single class of molecule. Using multiple markers on extracellular vesicles carry a huge potential for improving of understanding and managing of psychiatric disorders.