DAAO Human, Active

What is human D-amino acid oxidase and what is its primary function?

Human D-amino acid oxidase (hDAAO) is an FAD-containing flavoenzyme that catalyzes the oxidative deamination of D-amino acids with absolute stereoselectivity, with the exception of acidic D-amino acids. The reaction produces α-ketoacids, hydrogen peroxide, and ammonia. In humans, DAAO plays a critical role in degrading the neuromodulator D-serine in the central nervous system, where D-serine functions as the main coagonist of N-methyl-D-aspartate (NMDA) receptors . DAAO's activity must be finely tuned to fulfill this physiological function of controlling D-serine levels in the brain . The enzyme's structure is well conserved throughout evolution, with minor changes responsible for functional differences between microbial and human forms .

How does the FAD binding characteristic of human DAAO differ from other species?

Human DAAO possesses a uniquely weak interaction with its FAD cofactor, which is a distinguishing characteristic compared to DAAO from other species. Due to this weak interaction, human DAAO likely exists largely in the inactive apoprotein form in vivo . The binding of active-site ligands and substrates stabilizes flavin binding, promoting the acquisition of catalytic competence. This mechanism suggests that human DAAO activity can be regulated through the modulation of FAD binding, providing a control point for enzymatic function that may be distinct from DAAO regulation in other organisms .

What methodologies are commonly used to measure DAAO activity in laboratory settings?

Measurement of human DAAO activity typically employs fluorescence-based assays using the Amplex Red system. In this methodology, DAAO catalyzes the oxidative deamination of D-amino acids (commonly D-serine), producing hydrogen peroxide, which is then used by horseradish peroxidase (HRP) to oxidize Amplex Red to a fluorescent product. The fluorescent signal is measured with excitation at 530 nm and emission at 590 nm . Alternative methods include:

• Activity-based staining in tissue samples, which allows for the visualization of DAAO distribution in different brain regions

• Spectrophotometric assays that monitor the reduction of artificial electron acceptors

• Jump-dilution assays to study inhibitor-enzyme interaction kinetics

For accurate measurements, it's essential to include proper controls and consider the weak FAD binding characteristic of human DAAO by ensuring sufficient FAD concentration in the reaction mixture.

What is known about the substrate specificity of human DAAO?

Interestingly, in the nigrostriatal system, D-DOPA may be the preferential substrate of human DAAO instead of D-serine. DAAO efficiently metabolizes D-DOPA to dihydroxyphenylpyruvic acid, which can then be transaminated to L-DOPA through an alternative pathway for dopamine biosynthesis . This finding implies that DAAO may play a role in the metabolism of dopamine, norepinephrine, and epinephrine, extending its physiological significance beyond D-serine regulation.

How can researchers modulate the substrate specificity of human DAAO for experimental purposes?

Researchers can modulate DAAO substrate specificity through targeted mutations of residues located at the interface between the active site and the secondary binding pocket. By facilitating solvent and substrate access, it's possible to alter the enzyme's specificity profile . Specific approaches include:

• Identifying non-obvious residues on loops bordering the active site and secondary binding pocket that are essential for substrate specificity

• Engineering variants through site-directed mutagenesis of these critical residues

• Characterizing the biochemical activity of the resulting variants to confirm changes in specificity

Molecular dynamics simulations can complement structural analysis to characterize solvent accessibility and product release mechanisms before experimental validation . These approaches have successfully created DAAO variants with modified substrate preferences, which can be valuable for both industrial applications and studies of D-amino acid metabolism.

What kinetic mechanisms explain the inhibition patterns observed with human DAAO inhibitors?

Human DAAO inhibitors typically display an FAD-uncompetitive inhibition mechanism, similar to canonical inhibitors like benzoic acid . In this mechanism, inhibitors preferentially bind to the enzyme-FAD complex rather than to the apoenzyme. Understanding this mechanism is crucial for inhibitor design and experimentation.

To determine inhibitor mechanisms experimentally, researchers can perform saturation experiments using the Amplex Red system. By varying the concentration of either D-serine or FAD while keeping the other constant, and measuring enzyme activity in the presence of different inhibitor concentrations, the mode of inhibition can be established . Jump-dilution assays also provide valuable information about inhibitor binding kinetics, particularly the residence time of inhibitors on the enzyme.

These kinetic studies reveal that many DAAO inhibitors stabilize specific conformations of the enzyme, including a novel conformation where the active-site lid is in an open position, which has implications for structure-based drug design .

How does DAAO expression and activity vary across different human brain regions?

DAAO activity is widely distributed throughout the human central nervous system but follows specific patterns that differ significantly from those observed in mice. In humans, DAAO activity is detected in both forebrain and hindbrain regions, whereas in mice, it is predominantly hindbrain-shifted .

Specifically, in the human forebrain, DAAO activity is distributed in the subcortical white matter and the posterior limb of internal capsule, while it is almost undetectable in those areas in mice. Human DAAO activity is prominent along several neural pathways:

• Corticospinal tract

• Rubrospinal tract

• Nigrostriatal system

• Ponto-/olivo-cerebellar fibers

• Anterolateral system

In contrast, mice show strong DAAO activity primarily in the reticulospinal tract and ponto-/olivo-cerebellar fibers . This differential distribution may partly explain why DAAO-related conditions manifest differently between humans and rodent models.

What cellular and subcellular localization patterns does human DAAO exhibit in the central nervous system?

In the human central nervous system, DAAO activity does not colocalize with motoneuronal markers in the corticospinal tract but instead colocalizes predominantly with excitatory amino acid transporter 2 (EAAT2) and partially with glial fibrillary acidic protein (GFAP). This colocalization pattern suggests that DAAO activity-positive cells are primarily astrocytes, particularly those found in the motor pathway .

The subcellular localization of DAAO appears to be regulated through protein interactions. DAAO interacts with various proteins that modulate its activity, targeting, and cellular stability . This interaction network provides a mechanism for fine-tuning DAAO function at specific cellular locations.

Understanding the cellular and subcellular localization of DAAO is critical for interpreting its role in neurological disorders and for developing targeted therapeutic approaches. The predominant astrocytic localization in white matter tracts suggests roles beyond simple D-serine metabolism, possibly involving glial-neuronal signaling or maintenance of white matter integrity.

How does DAAO expression change during human brain development and aging?

DAAO expression undergoes dynamic changes during human brain development and aging. Studies examining normal human post-mortem brain samples ranging from 16 weeks of gestation to 91 years have revealed developmental regulation of both DAAO mRNA and protein expression across different brain regions .

The regulation of DAAO expression likely involves epigenetic mechanisms, including DNA methylation. DNA methylation patterns differ between cortical regions and the cerebellum in the human brain, potentially contributing to region-specific DAAO expression . Epigenetic regulation may play a crucial role in modulating DAAO levels during critical periods of brain development.

For researchers investigating DAAO's developmental profile, it's important to consider:

• Region-specific developmental trajectories

• Correlation between mRNA and protein levels

• Epigenetic marks associated with DAAO gene regulation

• Potential functional consequences of developmental changes in DAAO activity

These developmental patterns may contribute to understanding age-dependent susceptibility to neurological and psychiatric disorders in which DAAO has been implicated.

What evidence links DAAO to schizophrenia pathophysiology?

DAAO shows genetic association with schizophrenia, and abnormal D-serine metabolism has been implicated in the pathophysiology of this disorder . The glutamatergic hypofunction model of schizophrenia, which has gained significant support, suggests that reduced D-serine levels due to activation of DAAO may trigger hypofunction of NMDA receptors .

Since D-serine acts as a co-agonist at NMDA receptors, its degradation by DAAO could contribute to NMDA receptor hypofunction. This aligns with the glutamatergic hypothesis of schizophrenia, which proposes that NMDA receptor hypofunction leads to dysregulation of downstream dopaminergic systems .

The distribution of DAAO activity in the human brain, particularly in regions relevant to schizophrenia pathophysiology, provides anatomical support for this association. Researchers investigating the role of DAAO in schizophrenia should consider:

• The impact of DAAO genetic variants on enzyme activity and D-serine levels

• Region-specific effects of DAAO dysfunction in neural circuits implicated in schizophrenia

• Interactions between DAAO and other schizophrenia risk genes

• Potential therapeutic approaches targeting DAAO to enhance NMDA receptor function

How is DAAO implicated in amyotrophic lateral sclerosis (ALS) pathogenesis?

DAAO shows genetic association with amyotrophic lateral sclerosis (ALS), and its role in D-serine metabolism may contribute to disease pathophysiology . D-serine, as a co-agonist of NMDA receptors, can potentiate glutamatergic signaling. Excitotoxicity due to excessive glutamatergic stimulation is a well-established mechanism in ALS pathogenesis.

The distribution of DAAO activity in human brain motor pathways, particularly the corticospinal tract, provides anatomical evidence supporting its involvement in ALS . DAAO activity-positive cells in these pathways are primarily astrocytes, which are known to play critical roles in ALS progression.

For researchers investigating DAAO in ALS, important considerations include:

• The effects of ALS-associated DAAO variants on enzyme activity and D-serine metabolism

• Region-specific DAAO activity changes in ALS-affected brain regions

• The contribution of DAAO-mediated processes to motor neuron degeneration

• Potential therapeutic approaches targeting DAAO to modulate excitotoxicity in ALS

What methodological approaches can best evaluate DAAO's contribution to neuropsychiatric disorders?

Evaluating DAAO's contribution to neuropsychiatric disorders requires a multifaceted research approach:

• Genetic studies: Assessing associations between DAAO gene variants and disorder risk, considering both common and rare variants through sequencing and genome-wide association studies.

• Post-mortem brain analyses: Measuring DAAO mRNA, protein levels, and enzymatic activity in relevant brain regions from affected individuals and controls, using techniques like activity-based staining .

• Functional studies of disease-associated variants: Characterizing the biochemical properties of DAAO variants linked to disorders, including enzyme kinetics, substrate specificity, FAD binding, and protein-protein interactions .

• D-serine metabolism assessment: Measuring D-serine levels in cerebrospinal fluid or post-mortem brain tissue in relation to DAAO activity and disease status.

• Animal models: Developing transgenic models with human DAAO variants to evaluate behavioral and neurochemical phenotypes, while acknowledging species differences in DAAO distribution and function .

• Pharmacological studies: Testing selective DAAO inhibitors in disease models to establish causal relationships between DAAO activity and disease phenotypes.

• Circuit-specific analyses: Investigating DAAO's effects on specific neural circuits implicated in neuropsychiatric disorders using optogenetics or chemogenetics combined with DAAO manipulation.

These approaches should be integrated to overcome limitations of individual methods and to establish mechanistic links between DAAO dysfunction and disease pathophysiology.

How does the three-dimensional structure of human DAAO relate to its function?

The three-dimensional structure of human DAAO is well conserved throughout evolution, though subtle changes account for functional differences between microbial and human enzymes . The structure includes an active site where substrate binding and catalysis occur, a FAD-binding domain that accommodates the flavin cofactor, and dynamic regions such as the active-site lid that regulate substrate access.

Key structural features that influence function include:

• The active site architecture, which determines substrate specificity with absolute stereoselectivity for D-amino acids

• The FAD-binding pocket, characterized by relatively weak interactions with the flavin cofactor in human DAAO compared to other species

• The active-site lid, which can adopt different conformations (open or closed) affecting substrate access and product release

• Loops at the border between the active site and the secondary binding pocket that are critical for substrate specificity

Structural studies have revealed that human DAAO inhibitors can stabilize novel conformations, such as an open position of the active-site lid, confirming hypotheses about active-site lid flexibility . This structural information is valuable for structure-based drug design targeting human DAAO.

What strategies have been successful in engineering human DAAO for altered substrate specificity?

Engineering human DAAO for altered substrate specificity has been achieved through strategic modifications based on structural analysis and molecular dynamics simulations. Successful approaches include:

• Identification of key residues: Non-obvious residues located on loops at the border between the active site and secondary binding pocket have been identified as essential for substrate specificity .

• Facilitated solvent access: Mutations that modify solvent and substrate access to the active site have successfully modulated substrate specificity of both pig and human DAAO .

• Product egress pathway modification: Engineering residues involved in product release mechanisms has proven effective in altering enzyme activity and substrate preferences .

• Complementary computational and experimental approaches: Combining molecular dynamics simulations with biochemical characterization of enzyme variants has allowed for rational design of DAAO with modified specificity .

These engineering strategies have implications for both basic research, allowing investigation of structure-function relationships, and applied research, potentially enabling the development of DAAO variants with desired clinical or industrial properties.

What techniques are most effective for expressing and purifying active human DAAO for structural studies?

For structural studies of human DAAO, obtaining sufficient quantities of active, homogeneous enzyme is critical. The most effective techniques include:

• Expression systems: Recombinant expression in E. coli using hexaHistidine-tagged constructs has proven successful for both human and rat DAAO . For higher yields of soluble protein, expression at lower temperatures (15-20°C) after IPTG induction may be beneficial.

• Purification strategy:

  • Metal affinity chromatography using Ni-NTA resins for initial capture of His-tagged DAAO

  • Ion exchange chromatography for further purification

  • Size exclusion chromatography as a final polishing step to ensure homogeneity

• Stabilization considerations: Including FAD in purification buffers can enhance stability, though human DAAO has weak FAD binding. Adding glycerol (10-20%) and reducing agents can improve enzyme stability during purification.

• Activity preservation: Including competitive inhibitors during purification can sometimes protect the active site and enhance stability.

• Quality control: Assessing homogeneity by SDS-PAGE, measuring FAD content spectrophotometrically, and confirming enzymatic activity using standard assays are essential steps before proceeding to structural studies.

For crystallization specifically, preparing human DAAO in complex with inhibitors or substrate analogs has proven successful in stabilizing the enzyme and facilitating crystal formation . This approach has revealed novel conformations of the enzyme that are valuable for understanding its mechanism and for drug design.

How is human DAAO activity regulated at the cellular and molecular levels?

Human DAAO activity is regulated through multiple mechanisms at cellular and molecular levels:

Understanding these regulatory mechanisms is essential for interpreting DAAO's physiological roles and for developing strategies to modulate its activity in research or therapeutic contexts.

What is known about the role of DAOA/G72 in regulating human DAAO function?

DAOA (D-amino acid oxidase activator), also known as G72, is reported to interact with DAAO, but its effect on DAAO activity remains controversial and not fully understood. Research has yielded contradictory findings:

• Some studies report that DAOA increases DAAO activity , potentially enhancing D-serine degradation and thus affecting NMDA receptor function.

• Other investigations indicate that DAOA decreases DAAO activity , which would have the opposite effect on D-serine levels and NMDA receptor signaling.

• DAOA is localized in mitochondria and has been reported to modulate mitochondrial function , suggesting that its effects on DAAO may be part of broader cellular energy metabolism regulation.

The exact mechanism of DAOA-DAAO interaction and its physiological significance remain unclear. Both genes have been implicated in schizophrenia susceptibility, suggesting a potential role in the pathophysiology of this disorder through modulation of glutamatergic signaling.

For researchers investigating this interaction, important considerations include the expression patterns of both proteins, their subcellular localization, the conditions under which they interact, and the downstream consequences for D-serine metabolism and NMDA receptor function.

How do epigenetic mechanisms contribute to DAAO expression patterns in the human brain?

Epigenetic mechanisms appear to play a crucial role in regulating DAAO expression in the human brain, contributing to both regional specificity and developmental changes:

• DNA methylation patterns: DNA methylation levels in cortical regions differ from the cerebellum in the human brain , potentially contributing to region-specific DAAO expression. These methylation patterns can either increase or decrease gene expression, providing a flexible regulatory mechanism.

• Developmental regulation: Epigenetic modifications likely contribute to the dynamic changes in DAAO expression observed during brain development from gestation through aging . These modifications may establish developmental trajectories that determine DAAO activity in different brain regions throughout life.

• Region-specific regulation: The distinct patterns of DAAO expression and activity across brain regions may be maintained in part through epigenetic mechanisms that establish and preserve region-specific gene expression programs.

• Response to environmental factors: Epigenetic regulation provides a mechanism through which environmental factors might influence DAAO expression and activity, potentially contributing to individual variation in D-serine metabolism.

Understanding these epigenetic regulatory mechanisms is important not only for basic research on DAAO function but also for investigations of its role in neuropsychiatric disorders, where epigenetic dysregulation has been implicated.

What are the key considerations when designing experiments to measure human DAAO activity in tissue samples?

When designing experiments to measure human DAAO activity in tissue samples, researchers should consider:

• Tissue preservation: DAAO activity remains stable in post-mortem tissue for up to 48 hours , but rapid preservation is still preferable. Flash freezing in liquid nitrogen or immediate processing is recommended.

• Activity-based detection methods: For spatial distribution studies, activity-based staining methods can reveal DAAO activity patterns in tissue sections . These methods rely on the production of hydrogen peroxide during DAAO-catalyzed reactions.

• Specificity controls: Include D-amino acid substrate specificity controls and DAAO inhibitor controls to confirm the specificity of the measured activity.

• FAD supplementation: Due to the weak binding of FAD to human DAAO, ensuring sufficient FAD in reaction mixtures is crucial for accurate activity measurements.

• Regional variations: Consider the heterogeneous distribution of DAAO in the human brain when selecting regions for analysis . Include multiple regions to capture this variation.

• Cellular localization: Combine DAAO activity measurements with markers for specific cell types (e.g., GFAP for astrocytes, EAAT2 for excitatory amino acid transporters) to determine cellular localization .

• Quantification methods: For biochemical assays, fluorometric methods using Amplex Red are sensitive for detecting hydrogen peroxide production . Ensure linear range of detection and appropriate controls.

• Species differences: Be aware that findings from rodent models may not translate directly to humans due to significant differences in DAAO distribution and activity levels between species .

These considerations will help ensure reliable and interpretable measurements of human DAAO activity in tissue samples for both basic research and clinical investigations.

How can researchers effectively study the interaction between human DAAO and its inhibitors?

To effectively study interactions between human DAAO and its inhibitors, researchers can employ several complementary approaches:

• Enzyme kinetic assays: Determine inhibition mechanisms (competitive, uncompetitive, or noncompetitive) by measuring enzyme activity with varying substrate and inhibitor concentrations. Many DAAO inhibitors show FAD-uncompetitive inhibition .

• Jump-dilution assays: Assess inhibitor binding kinetics and residence time by pre-incubating enzyme with inhibitor at high concentration, then diluting into substrate-containing solution and monitoring activity recovery over time .

• Thermal shift assays: Measure changes in protein thermal stability upon inhibitor binding, which can indicate binding affinity and conformational effects.

• X-ray crystallography: Co-crystallize DAAO with inhibitors to determine binding modes and structural changes. This approach has revealed that inhibitors can stabilize novel conformations of DAAO, including an open position of the active-site lid .

• Molecular dynamics simulations: Complement experimental studies with computational analyses to understand inhibitor binding dynamics and effects on protein conformation.

• Cellular assays: Evaluate inhibitor effects in cellular contexts where DAAO is expressed, measuring changes in D-amino acid levels or downstream signaling.

• Structure-activity relationship studies: Systematically modify inhibitor structures and correlate with binding affinity and inhibitory potency to understand critical interaction features.

These approaches provide comprehensive insights into inhibitor mechanisms, binding modes, and potential applications in research or therapeutic development.

What experimental approaches best address the species differences in DAAO distribution and function?

Addressing species differences in DAAO distribution and function requires specialized experimental approaches:

• Comparative activity mapping: Apply identical activity-based staining methods to human and rodent brain samples to directly compare regional distribution patterns . This approach has revealed that human DAAO activity is distributed in the subcortical white matter and internal capsule, whereas it is almost undetectable in these areas in mice.

• Cross-species biochemical characterization: Compare enzymatic properties (substrate specificity, kinetic parameters, FAD binding) of purified human and rodent DAAO under identical conditions to identify functional differences.

• Humanized animal models: Generate transgenic mice expressing human DAAO under native regulatory elements to better model human-specific aspects of DAAO function in vivo.

• Cell type-specific analysis: Combine DAAO activity measurements with cell type-specific markers in both human and rodent tissues to determine whether cellular localization differs between species . For example, in humans, DAAO activity-positive cells in the corticospinal tract are primarily astrocytes.

• Pathway-specific approaches: Focus on specific neural pathways (e.g., corticospinal tract, nigrostriatal system) when comparing species, as DAAO distribution follows different pathway patterns in humans versus rodents .

• Cross-species transcriptomics and proteomics: Compare DAAO expression patterns and protein interaction networks between species to identify regulatory differences.

• Human cell models: Utilize human cell models (primary cultures, iPSC-derived neural cells) to study DAAO regulation and function in a human cellular context.

These approaches help translate findings between animal models and humans, crucial for understanding DAAO's role in human neurological disorders and for developing therapeutics targeting this enzyme.

What are the most promising therapeutic applications of DAAO modulation in neurological disorders?

Modulation of DAAO activity holds promise for several therapeutic applications in neurological disorders:

• Schizophrenia treatment: Inhibiting DAAO to increase D-serine levels could enhance NMDA receptor function, potentially addressing the glutamatergic hypofunction implicated in schizophrenia pathophysiology . This approach offers an alternative to direct D-serine supplementation, which faces pharmacokinetic challenges.

• Amyotrophic lateral sclerosis (ALS): Targeting DAAO in motor pathways, where it shows prominent activity in humans , could modulate excitotoxicity contributing to motor neuron degeneration in ALS.

• Pain management: Given DAAO's role in D-amino acid metabolism and its relevance to neuropathic pain , selective DAAO inhibitors could represent a novel approach to pain management without the addictive potential of current treatments.

• Precision medicine approaches: Genetic variants of DAAO associated with neuropsychiatric disorders could guide personalized therapeutic strategies, targeting specific dysfunctional aspects of D-serine metabolism in individual patients.

• Combination therapies: DAAO inhibition could potentially enhance the efficacy of existing treatments for neurological disorders by modulating glutamatergic signaling through increased D-serine availability.

Future research should focus on developing highly selective DAAO inhibitors with favorable pharmacokinetic properties, understanding region-specific effects of DAAO modulation, and identifying patient populations most likely to benefit from DAAO-targeted therapies.

What unresolved questions remain regarding the physiological role of DAAO in humans?

Despite significant advances, several crucial questions about DAAO's physiological role in humans remain unresolved:

• Substrate preference in different brain regions: While D-serine is considered DAAO's primary substrate in most brain regions, evidence suggests that D-DOPA might be the preferential substrate in the nigrostriatal system . The region-specific substrate preferences and their physiological significance require further investigation.

• Function in white matter tracts: DAAO activity is prominent along several white matter tracts in humans , but its function in these regions is poorly understood. Does it contribute to axonal integrity, myelin maintenance, or glial-neuronal signaling?

• Developmental roles: The developmental regulation of DAAO expression suggests potential roles during brain development, but these functions remain largely unexplored.

• Regulation of enzymatic activity: Given the weak FAD binding of human DAAO , what mechanisms ensure appropriate enzyme activity in vivo? How do protein interactions, post-translational modifications, and subcellular targeting regulate DAAO function?

• Contribution to non-neurological functions: DAAO's role outside the nervous system, including potential functions in immune regulation or metabolic processes, remains understudied.

• Evolutionary significance: Why has human DAAO evolved distinct characteristics compared to other species, including broader brain distribution and weaker FAD binding? What selective pressures drove these changes?

Addressing these questions will require integrated approaches combining biochemical, cellular, and systems-level investigations in human tissues and model systems.

How might advanced technologies enhance our understanding of DAAO in human neurological health and disease?

Advanced technologies offer exciting opportunities to deepen our understanding of DAAO in human neurological health and disease:

• Single-cell omics: Single-cell RNA sequencing and proteomics can reveal cell type-specific expression patterns of DAAO and its regulatory partners across brain regions, providing insights into its functional heterogeneity.

• CRISPR-based approaches: Gene editing in human cell models can help evaluate the functional consequences of DAAO variants identified in neurological disorders and enable high-throughput screening of genetic modifiers.

• Advanced imaging techniques: Super-resolution microscopy and expansion microscopy can visualize DAAO's subcellular localization and its proximity to interaction partners with unprecedented detail.

• Cryo-electron microscopy: This technique can capture dynamic conformational states of DAAO that are challenging to observe by X-ray crystallography, potentially revealing new regulatory mechanisms.

• In vivo D-serine sensors: Development of genetically encoded sensors for D-serine would allow real-time monitoring of DAAO activity consequences in living systems.

• Brain organoids: Human brain organoids provide three-dimensional models to study DAAO function in a complex cellular environment that recapitulates aspects of human brain development.

• Computational approaches: Machine learning algorithms applied to large datasets could identify novel patterns in DAAO expression, regulation, and association with disease phenotypes.

• Spatial transcriptomics and proteomics: These techniques can map DAAO expression and activity in relation to other genes and proteins across intact brain regions, revealing functional networks.

Integration of these advanced technologies promises to transform our understanding of DAAO's complex roles in human neurological health and disease, potentially leading to novel therapeutic strategies.