Final Report Summary - SLEEP PLASTICITY (The role of rhythmic synaptic plasticity in regulating sleep and behavioral performance)
Sleep is an essential biological need of higher organisms. The functions of sleep are debated and include memory consolidation and synaptic plasticity. To understand the molecular mechanisms that regulate structural synaptic plasticity and sleep, we used the zebrafish model. The zebrafish is a diurnal transparent vertebrate with high-throughput genetics that is suitable for time-lapse two-photon imaging of pre- and post-synaptic markers in live animal. The goal of this proposal was to identify and characterize new synaptic proteins and functional circuits required for sleep and circadian clock regulation. In order to achieve this goal, we formulated the following objectives: 1. to identify and characterize brain regions demonstrating rhythmic and sleep-dependent synaptic plasticity; and 2. to discover the functional effect of circadian and sleep-dependent synaptic changes on rhythmic behavior, such as sleep.
In the last four years, the results were published in six papers:
In the hypothalamus, we described the function of hypocretin/orexin (HCRT) neurons and identified a novel neurotensin (NTS) secreting neurons. We established an HCRT-neuron–ablated zebrafish as a model for narcolepsy. This fish exhibited an increase in sleep and sleep/wake transitions, as well as altered response to external light and sound stimuli. These results were published in the Journal of Neuroscience in 2012 (Elbaz et al. 2012). The role of HCRT neurons in regulating sleep and the description of live imaging of synaptic marker during day and night were summarized in a review that published by us in Frontiers in Neural Circuits (Elbaz et al. 2013). In addition, we cloned the nts gene and promoter and show interaction between NTS secreting neurons and the HCRT system. This data was published by us in the Journal of Comparitve Neurology (Levitas-Djerbi et al. 2015).
We also used behavioral systems and live imaging to link synaptic density with sleep deficiencies in zebrafish model for mental retardation. We generated an mct8 mutant (mct8-/-) zebrafish using zinc-finger nuclease (ZFN)-mediated targeted gene editing system. Lack of MCT8 in human disturbs thyroid hormone levels and result in mental retardation. The mct8-/- larvae showed reduced synaptic density in several neuronal circuits and increased sleep. These results were published by us in two journals: the Journal of Biological Chemistry (Vatine et al. 2013) and PLoS genetics (Zada et al. 2014).
In order to study rhythmic structural synaptic plasticity we characterized the expression of nptx2a and established a transcription activator-like effector nuclease (TALEN)–mediated nptx2a mutant (nptx2a-/-) zebrafish. Behavioral assays showed that loss of Nptx2a results in reduced locomotor response to light-to-dark transition states and to a sound stimulus. Live imaging of synapses using the transgenic nptx2a:GAL4VP16 zebrafish and a fluorescent presynaptic synaptophysin (SYP) marker revealed reduced synaptic density in the axons of the spinal motor neurons and the anterodorsal lateral-line ganglion (gAD), which regulate locomotor activity and locomotor response to mechanosensory stimuli, respectively. Furthermore, the number of synapses in the gAD was rhythmic during day and night. These results suggest that Nptx2a affects locomotor response to external stimuli by mediating structural synaptic plasticity in excitatory neuronal circuits. This work was published by us in FASEB journal (Elbaz et al. 2015).
Altogether, we established several fish model for sleep disorders and characterized the function of genes and neuronal circuits that regulate sleep. We also visualized rhythmic structural synaptic plasticity in several neuronal circuits of live vertebrate.
In the last four years, the results were published in six papers:
In the hypothalamus, we described the function of hypocretin/orexin (HCRT) neurons and identified a novel neurotensin (NTS) secreting neurons. We established an HCRT-neuron–ablated zebrafish as a model for narcolepsy. This fish exhibited an increase in sleep and sleep/wake transitions, as well as altered response to external light and sound stimuli. These results were published in the Journal of Neuroscience in 2012 (Elbaz et al. 2012). The role of HCRT neurons in regulating sleep and the description of live imaging of synaptic marker during day and night were summarized in a review that published by us in Frontiers in Neural Circuits (Elbaz et al. 2013). In addition, we cloned the nts gene and promoter and show interaction between NTS secreting neurons and the HCRT system. This data was published by us in the Journal of Comparitve Neurology (Levitas-Djerbi et al. 2015).
We also used behavioral systems and live imaging to link synaptic density with sleep deficiencies in zebrafish model for mental retardation. We generated an mct8 mutant (mct8-/-) zebrafish using zinc-finger nuclease (ZFN)-mediated targeted gene editing system. Lack of MCT8 in human disturbs thyroid hormone levels and result in mental retardation. The mct8-/- larvae showed reduced synaptic density in several neuronal circuits and increased sleep. These results were published by us in two journals: the Journal of Biological Chemistry (Vatine et al. 2013) and PLoS genetics (Zada et al. 2014).
In order to study rhythmic structural synaptic plasticity we characterized the expression of nptx2a and established a transcription activator-like effector nuclease (TALEN)–mediated nptx2a mutant (nptx2a-/-) zebrafish. Behavioral assays showed that loss of Nptx2a results in reduced locomotor response to light-to-dark transition states and to a sound stimulus. Live imaging of synapses using the transgenic nptx2a:GAL4VP16 zebrafish and a fluorescent presynaptic synaptophysin (SYP) marker revealed reduced synaptic density in the axons of the spinal motor neurons and the anterodorsal lateral-line ganglion (gAD), which regulate locomotor activity and locomotor response to mechanosensory stimuli, respectively. Furthermore, the number of synapses in the gAD was rhythmic during day and night. These results suggest that Nptx2a affects locomotor response to external stimuli by mediating structural synaptic plasticity in excitatory neuronal circuits. This work was published by us in FASEB journal (Elbaz et al. 2015).
Altogether, we established several fish model for sleep disorders and characterized the function of genes and neuronal circuits that regulate sleep. We also visualized rhythmic structural synaptic plasticity in several neuronal circuits of live vertebrate.