Synapses are essential components of neurons and allow information to travel coordinately throughout the nervous system to adjust behavior to environmental stimuli and to control body functions, memories, and emotions. Thus, optimal synaptic communication is required for proper brain physiology, and slight perturbations of synapse function can lead to brain disorders. In fact, increasing evidence has demonstrated the relevance of synapse dysfunction as a major determinant of many neurological diseases. This notion has led to the concept of synaptopathies as brain diseases with synapse defects as shared pathogenic features. In this review, which was initiated at the 13th International Society for Neurochemistry Advanced School, we discuss basic concepts of synapse structure and function, and provide a critical view of how aberrant synapse physiology may contribute to neurodevelopmental disorders (autism, Down syndrome, startle disease, and epilepsy) as well as neurodegenerative disorders (Alzheimer and Parkinson disease). We finally discuss the appropriateness and potential implications of gathering synapse diseases under a single term. Understanding common causes and intrinsic differences in diseaseassociated synaptic dysfunction could offer novel clues toward synapse-based therapeutic intervention for neurological and neuropsychiatric disorders.

Recent studies on the pathogenic mechanisms of recessive hyperekplexia indicate disturbances in glycine receptor (GlyR) ␣1 biogenesis. Here, we examine the properties of a range of novel glycine receptor mutants identified in human hyperekplexia patients using expression in transfected cell lines and primary neurons. All of the novel mutants localized in the large extracellular domain of the GlyR ␣1 have reduced cell surface expression with a high proportion of receptors being retained in the ER, although there is forward trafficking of glycosylated subpopulations into the ER-Golgi intermediate compartment and cis-Golgi compartment. CD spectroscopy revealed that the mutant receptors have proportions of secondary structural elements similar to wild-type receptors. Two mutants in loop B (G160R, T162M) were functional, but none of those in loop D/␤2-3 were. One nonfunctional truncated mutant (R316X) could be rescued by coexpression with the lacking C-terminal domain. We conclude that a proportion of GlyR ␣1 mutants can be transported to the plasma membrane but do not necessarily form functional ion channels. We suggest that loop D/␤2-3 is an important determinant for GlyR trafficking and functionality, whereas alterations to loop B alter agonist potencies, indicating that residues here are critical elements in ligand binding.

Functional impairments or trafficking defects of inhibitory glycine receptors (GlyRs) have been linked to human hyperekplexia/startle disease and autism spectrum disorders. We found that a lack of synaptic integration of GlyRs, together with disrupted receptor function, is responsible for a lethal startle phenotype in a novel spontaneous mouse mutant shaky, caused by a missense mutation, Q177K, located in the extracellular β8–β9 loop of the GlyR α1 subunit. Recently, structural data provided evidence that the flexibility of the β8–β9 loop is crucial for conformational transitions during opening and closing of the ion channel and represents a novel allosteric binding site in Cys-loop receptors. We identified the underlying neuropathological mechanisms in male and female shaky mice through a combination of protein biochemistry, immunocytochemistry, and both in vivo and in vitro electrophysiology. Increased expression of the mutant GlyR α1Q177K subunit in vivo was not sufficient to compensate for a decrease in synaptic integration of α1Q177Kβ GlyRs. The remaining synaptic heteromeric α1Q177Kβ GlyRs had decreased current amplitudes with significantly faster decay times. This functional disruption reveals an important role for the GlyR α1 subunit β8–β9 loop in initiating rearrangements within the extracellular–transmembrane GlyR interface and that this structural element is vital for inhibitory GlyR function, signaling, and synaptic clustering.SIGNIFICANCE STATEMENT GlyR dysfunction underlies neuromotor deficits in startle disease and autism spectrum disorders. We describe an extracellular GlyR α1 subunit mutation (Q177K) in a novel mouse startle disease mutant shaky. Structural data suggest that during signal transduction, large transitions of the β8–β9 loop occur in response to neurotransmitter binding. Disruption of the β8–β9 loop by the Q177K mutation results in a disruption of hydrogen bonds between Q177 and the ligand-binding residue R65. Functionally, the Q177K change resulted in decreased current amplitudes, altered desensitization decay time constants, and reduced GlyR clustering and synaptic strength. The GlyR β8–β9 loop is therefore an essential regulator of conformational rearrangements during ion channel opening and closing.

The molecular genetics of panic disorder (PD) with and without agoraphobia (AG) are still largely unknown and progress is hampered by small sample sizes. We therefore performed a genome-wide association study with a dimensional, PD/AG -related anxiety phenotype based on the Agoraphobia Cognition Questionnaire (ACQ) in a sample of 1,370 healthy German volunteers of the CRC TRR58 MEGA study wave 1. A genome-wide significant association was found between ACQ and single non-coding nucleotide variants of the GLRB gene (rs78726293, p=3.3x10 -8 ; rs191260602, p=3.9x10 -8 ). We followed up on this finding in a larger dimensional ACQ sample (N=2,547) and in independent samples with a dichotomous AG phenotype based on the Symptoms Checklist (SCL-90; N=3,845) and a casecontrol sample with the categorical phenotype PD/AG (N combined =1,012) obtaining highly Partial Glrb knockout-mice demonstrated an agoraphobic phenotype. In conjunction with the clinical observation that rare coding GLRB gene mutations are associated with the neurological disorder hyperekplexia characterized by a generalized startle reaction and agoraphobic behavior, our data provide evidence that non-coding, though functional GLRB gene polymorphisms may predispose to PD by increasing startle response and agoraphobic cognitions.

Highlights d First structure of the anti-malarial drug artemisinin bound to a target protein d Artemisinins target the glycine and GABA A receptor binding pocket in gephyrin d Artemisinins modulate inhibitory neurotransmission with a dependence on gephyrin d Gephyrin-receptor clusters are targeted by artemisinins in a time-dependent manner

In this study, we designed a novel biomaterial ink platform based on hydrophilic poly(2-ethyl-2-oxazine) (PEtOzi) specifically for melt electrowriting (MEW). This material crosslinks spontaneously after processing via dynamic Diels-Alder click chemistry. These direct-written microperiodic structures rapidly swell in water to yield thermoreversible hydrogels. These hydrogels are robust enough for repeated aspiration and ejection through a cannula without structural damage, despite their high water content of 84%. Moreover, the scaffolds retain functional groups for modification using click chemistry and therefore can be readily functionalized as demonstrated using fluorophores and peptides to facilitate visualization and cell attachment. The PEtOzi hydrogel developed here is compatible with confocal imaging and staining protocols for cells. In summary, an advanced material platform based on PEtOzi is reported that is compatible with MEW and results in functionalizable chemically crosslinked microperiodic hydrogels.

Ionotropic glycine receptors (GlyRs) enable fast synaptic neurotransmission in the adult spinal cord and brainstem. The inhibitory GlyR is a transmembrane glycine-gated chloride channel. The immature GlyR protein undergoes various processing steps, e.g., folding, assembly, and maturation while traveling from the endoplasmic reticulum to and through the Golgi apparatus, where post-translational modifications, e.g., glycosylation occur. The mature receptors are forward transported via microtubules to the cellular surface and inserted into neuronal membranes followed by synaptic clustering. The normal life cycle of a receptor protein includes further processes like internalization, recycling, and degradation. Defects in GlyR life cycle, e.g., impaired protein maturation and degradation have been demonstrated to underlie pathological mechanisms of various neurological diseases. The neurological disorder startle disease is caused by glycinergic dysfunction mainly due to missense mutations in genes encoding GlyR subunits (GLRA1 and GLRB). In vitro studies have shown that most recessive forms of startle disease are associated with impaired receptor biogenesis. Another neurological disease with a phenotype similar to startle disease is a special form of stiff-person syndrome (SPS), which is most probably due to the development of GlyR autoantibodies. Binding of GlyR autoantibodies leads to enhanced receptor internalization. Here we focus on the normal life cycle of GlyRs concentrating on assembly and maturation, receptor trafficking, post-synaptic integration and clustering, and GlyR internalization/recycling/degradation. Furthermore, this review highlights findings on impairment of these processes under disease conditions such as disturbed neuronal ER-Golgi trafficking as the major pathomechanism for recessive forms of human startle disease. In SPS, enhanced receptor internalization upon autoantibody binding to the GlyR has been shown to underlie the human pathology. In addition, we discuss how the existing mouse models of startle disease increased our current knowledge of GlyR trafficking routes and function. This review further illuminates receptor trafficking of GlyR variants originally identified in startle disease patients and explains changes in the life cycle of GlyRs in patients with SPS with respect to structural and functional consequences at the receptor level.

of the matrix. [9] Mechanical properties such as the stiffness of the 3D surrounding environment are known to affect differentiation of certain cells, [10] and are believed to be critical for neuronal maturation and neurotransmission. [9] More discrete and higher resolved structures, such as those made with additive manufacturing (AM) [11] and electrojetting technology, [12] are promising to provide reproducible conditions. Electrojetting uses electrostatic forces to fabricate monodispersed, nanomicro particles in a simple, versatile, and cost-effective method for drug delivery and tissue engineering applications. [13] Electrospinning allows processability for polymer solutions and polymer melts. [14] In addition, polymer melts can be directly written using a programmable path in a technique known as melt electrowriting (MEW). [15] The precise placement of low-micrometer diameter fibers that are stackable using additive manufacturing principles are achievable using MEW (Figure 1a,b). The advantages of MEW include solvent-free processing and avoidance of the chaotic fiber deposition often seen in electrospinning. Bioprinting approaches for cell hierarchy have been reinforced with extruded, stiffer structures [16] that reinforce the bioink and aid with in vitro handling [17] and extend a processing window for such hydrogels. [16,17] The reinforcing of matrices and bioinks with much smaller, low-micrometer scale fibers on matrices has recently been of particular interest. [18,19] Using MEW, [15] well-ordered, small diameter fibers can be distributed throughout a matrix in low volume fractions and provide significant increase in overall mechanical properties. [18][19][20] Furthermore, the mechanics of MEW fiber-reinforced hydrogels can be modulated further with sinusoidal direct-writing of the fibers. [18] Since the fiber diameter made using MEW can be readily changed with the nozzle mass flow rate and/or the collection speed, the diameter of the printed fiber can be significantly altered. [21] Therefore, MEW reinforcement of matrices (such as Matrigel shown here) have the potential to regulate the environment of cells through both mechanotransductive [22] or haptotaptic [23] cues.An experimentally designed in vitro 3D structure for electrophysiological studies requires a relevant receptor model, with the inhibitory glycine receptor (GlyR) used in this study. The GlyR is a pentameric, ligand gated ion channel, which belongs to the superfamily of Cys-loop receptors. [24] Upon binding of 2D electrophysiology is often used to determine the electrical properties of neurons. In the brain however, neurons form extensive 3D networks. Thus, performing electrophysiology in a 3D environment provides a closer situation to the physiological condition and serves as a useful tool for various applications in the field of neuroscience. In this study, 3D electrophysiology is established within a fiber-reinforced matrix to enable fast readouts from transfected cells, which are often used as model systems for 2D electrophysiology. Usin...