|dc.description.abstracteng||During early postnatal life, the brain goes through critical periods of increased plasticity, which enable experience-dependent refinements of neuronal circuits. Insertion of α-amino- 3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPAR) to the post-synaptic membrane of nascent AMPAR-silent synapses during the critical periods is an essential neurodevelopmental process, enabling experience-dependent strengthening of favored glutamatergic connections. One of the key molecules that promotes AMPAR-silent synapse maturation is postsynaptic density (PSD) protein-95. It was recently shown that deleting PSD-95 halts the silent-synapse maturation at a developmentally immature state and prevents the closure of critical period for ocular dominance plasticity (ODP), a classical paradigm for studying the cortical plasticity (Wiesel and Hubel, 1963b). In adult mice lacking PSD-95, monocularly depriving the contralateral eye of the visual input for 4 days leads to a strong reduction in deprived-eye responses in the binocular part of the primary visual cortex (V1), and the plastic changes are reversed within 2 days of reopening the eye (Huang et al., 2015)—a phenotype classically seen only in juvenile wild-type mice if reared in standard conditions (Espinosa and Stryker, 2012).
In order to understand the underlying morphological correlates of enhanced cortical plasticity in the absence of PSD-95, I set out to investigate experience-dependent dendritic spine dynamics in layer 2/3 pyramidal neurons of binocular V1 in awake PSD-95 knockout (KO) and knockdown (KD) mice using awake, in-vivo two-photon imaging. Layer 2/3 pyramidal neurons in V1 were selectively transfected to express green fluorescent protein using in utero electroporation at embryonic day 15.5. Surgically implanting a long-term cranial window over V1 enabled repeated optical access to the superficial layers of the cortex. I imaged the same set of apical dendrites of layer 2/3 pyramidal neurons repeatedly, during and after a 4-day monocular deprivation (MD) of the contralateral eye in postnatal day (P) ~75 animals. Analyses of spine formation and elimination rates during these phases revealed that in PSD-95 KO mice, 4-day MD perturbs spine dynamics towards significantly increased spine elimination and a relative decrease in spine formation, while in WT controls spine dynamics remained essentially unchanged. The MD-induced imbalance in spine dynamics led to a significant reduction in spine density in PSD-95 KO mice. Interestingly, although the percentage of persistent spines was similar in KO and WT mice over the days of imaging, in PSD-95 KO mice newly formed spines were significantly more likely to be eliminated after MD. During the subsequent recovery phase, spine elimination ratio remained elevated in PSD-95 KO mice compared to baseline, although it was not anymore significantly different compared to WT controls. Similarly, in PSD-95 KD neurons MD induced a significant increase in spine elimination compared to controls, consistent with cell-autonomous impact of PSD-95 on AMPAR-silent synapse fractions. Overall, in PSD- 95-deficient mice, MD-dependent changes on spine dynamics were similar to that of previously described in juvenile mice (Sun et al., 2019), suggesting that absence of PSD- 95 results in juvenile-like plastic state.
Neurogranin, a postsynaptic signaling protein, was recently shown to also promote the AMPAR-silent synapse maturation and experience-dependent spine elimination during the time frame of critical period for ODP (Han et al., 2017). However, its role in critical period closure had not been investigated. Due to the elevated fraction of AMPAR-silent synapses in neurogranin-deficient mice, we hypothesized that the critical period for ODP should also remain open in these mice, similar to what has been observed previously in PSD-95- deficient mice (Huang et al., 2015). Using optical imaging of intrinsic signals to assess ODP, here I show that knockdown of neurogranin at birth indeed i) prevented the closure of the critical period for ODP as observed for standard-cage raised mice (Gordon and Stryker, 1996), and ii) enhanced ODP even in mice up to at least P149, copying the phenotype previously observed in PSD-95-deficient mice. Moreover, while innate visual capabilities of adult neurogranin knockdown mice remained intact, their visual acuity was impaired, which is phenotypically different from PSD-95 deficient mice (Han et al., 2017; Huang et al., 2015). Thus, neurogranin is required for both the closure of the critical period for ODP and the functional development of V1 circuitry for optimal performance.
Overall, the findings in this thesis further substantiate the idea that enhanced brain plasticity during the early life is closely linked to the availability of AMPAR-silent synapses. Perturbing the molecules regulating the AMPAR-silent synapse maturation in the primary visual cortex, confers the cortical circuitry juvenile-like ODP plasticity, both structurally and physiologically.||de