Altered Behavioral Performance and Live Imaging of Circuit-Specific Neural Deficiencies in a Zebrafish Model for Psychomotor Retardation

Abstract

The mechanisms and treatment of psychomotor retardation, which includes motor and cognitive impairment, are indefinite. The Allan-Herndon-Dudley syndrome (AHDS) is an X-linked psychomotor retardation characterized by delayed development, severe intellectual disability, muscle hypotonia, and spastic paraplegia, in combination with disturbed thyroid hormone (TH) parameters. AHDS has been associated with mutations in the monocarboxylate transporter 8 (mct8/slc16a2) gene, which is a TH transporter. In order to determine the pathophysiological mechanisms of AHDS, MCT8 knockout mice were intensively studied. Although these mice faithfully replicated the abnormal serum TH levels, they failed to exhibit the neurological and behavioral symptoms of AHDS patients. Here, we generated an mct8 mutant (mct8−/−) zebrafish using zinc-finger nuclease (ZFN)-mediated targeted gene editing system. The elimination of MCT8 decreased the expression levels of TH receptors; however, it did not affect the expression of other TH-related genes. Similar to human patients, mct8−/− larvae exhibited neurological and behavioral deficiencies. High-throughput behavioral assays demonstrated that mct8−/− larvae exhibited reduced locomotor activity, altered response to external light and dark transitions and an increase in sleep time. These deficiencies in behavioral performance were associated with altered expression of myelin-related genes and neuron-specific deficiencies in circuit formation. Time-lapse imaging of single-axon arbors and synapses in live mct8−/− larvae revealed a reduction in filopodia dynamics and axon branching in sensory neurons and decreased synaptic density in motor neurons. These phenotypes enable assessment of the therapeutic potential of three TH analogs that can enter the cells in the absence of MCT8. The TH analogs restored the myelin and axon outgrowth deficiencies in mct8−/− larvae. These findings suggest a mechanism by which MCT8 regulates neural circuit assembly, ultimately mediating sensory and motor control of behavioral performance. We also propose that the administration of TH analogs early during embryo development can specifically reduce neurological damage in AHDS patients.

Sleep and Development in Genetically Tractable Model Organisms

“…These fly models of neurodevelopmental disorders provide an opportunity to dissect sleep by development interactions with the goal of a deeper genetic/molecular understanding of sleep in neurodevelopmental disorders. Of note, this line of work will not be limited to the fly going forward: a zebrafish model of AllanHerndon-Dudley syndrome (ADHS) exhibits both abnormal (high) sleep levels and responses to light-dark transitions (Zada et al 2014), and it is likely that additional models will continue to emerge. There are multiple questions uniquely suited to examination in simple genetic systems: Are specific sleep/circadian neural loci perturbed in neurodevelopmental disorders?…”

MCT8 deficiency in Purkinje cells disrupts embryonic chicken cerebellar development

“…Despite the generation and extensive study of different MCT8-deficient animal models (Dumitrescu et al 2006, de Vrieze et al 2014, Zada et al 2014, the exact pathway by which MCT8 regulates cerebellar circuit organisation remains unclear. Mct8-deficient zebrafish display an underdeveloped cerebellum and exhibit clear locomotor deficits (de Vrieze et al 2014, Zada et al 2014.…”

“…Mct8-deficient zebrafish display an underdeveloped cerebellum and exhibit clear locomotor deficits (de Vrieze et al 2014, Zada et al 2014. In mice, however, apart from phenocopying the altered serum TH profile, Mct8 knockout (KO) results in an apparently unaffected cerebellar structure (Trajkovic et al 2007, Koibuchi 2009, coupled with normal performance in locomotor behavioural tests (Wirth et al 2009).…”

“…Although some of them were shown to have beneficial effects in Mct8 KO zebrafish larvae (Zada et al 2014) and in Mct8/Oatp1c1 double KO mice (Horn et al 2013, Kersseboom et al 2014, a recent clinical trial in patients treated in the first year of infancy failed to improve mental status, probably because neurological damage was already irreversible at the onset of treatment (Verge et al 2012). A better understanding of the function of MCT8 at the neuronal level during embryonic development is therefore an important objective.…”

Zebrafish Models of Human Disease: Gaining Insight into Human Disease at ZFIN

“…The improvement of reverse genetic techniques such as the use of zinc finger nucleases (Leong et al 2011; Sander et al 2011a, 2011b) CRISPRs (Hruscha and Schmid 2015; Hwang et al 2013; Hwang et al 2015) and transcription activator-like effector nucleases (TALENs; Huang et al 2011, 2016; Ma et al 2016; Sander et al 2011a, 2011b) have allowed for the targeted mutation of disease-specific genes. These techniques are becoming the method of choice in zebrafish for creating genetic models of human disease, such as models for visceral heterotaxy, Bethlem myopathy, and Allan-Herndon-Dudley syndrome (Table 1A; Noël et al 2015; Radev et al 2015; Zada et al 2014). …”