Faculty Bibliography

BACKGROUND: Alzheimer's disease (AD) is characterized by amyloid deposition, tangle formation as well as synapse loss. Synaptic abnormalities occur early in the pathogenesis of AD. Identifying early synaptic abnormalities and their underlying mechanisms is likely important for the prevention and treatment of AD. METHODS: We performed in vivo two-photon calcium imaging to examine the activities of somas, dendrites and dendritic spines of layer 2/3 pyramidal neurons in the primary motor cortex in the APPswe/PS1dE9 mouse model of AD and age-matched wild type control mice. We also performed calcium imaging to determine the effect of Abeta oligomers on dendritic calcium activity. In addition, structural and functional two-photon imaging were used to examine the link between abnormal dendritic calcium activity and changes in dendritic spine size in the AD mouse model. RESULTS: We found that somatic calcium activities of layer 2/3 neurons were significantly lower in the primary motor cortex of 3-month-old APPswe/PS1dE9 mice than in wild type mice during quiet resting, but not during running on a treadmill. Notably, a significantly larger fraction of apical dendrites of layer 2/3 pyramidal neurons showed calcium transients with abnormally long duration and high peak amplitudes during treadmill running in AD mice. Administration of Abeta oligomers into the brain of wild type mice also induced abnormal dendritic calcium transients during running. Furthermore, we found that the activity and size of dendritic spines were significantly reduced on dendritic branches with abnormally prolonged dendritic calcium transients in AD mice. CONCLUSION: Our findings show that abnormal dendritic calcium transients and synaptic depotentiation occur before amyloid plaque formation in the motor cortex of the APPswe/PS1dE9 mouse model of AD. Dendritic calcium transients with abnormally long durations and high amplitudes could be induced by soluble Abeta oligomers and contribute to synaptic deficits in the early pathogenesis of AD.

Neuropathic pain involves long-lasting modifications of pain pathways that result in abnormal cortical activity. How cortical circuits are altered and contribute to the intense sensation associated with allodynia is unclear. Here we report a persistent elevation of layer V pyramidal neuron activity in the somatosensory cortex of a mouse model of neuropathic pain. This enhanced pyramidal neuron activity was caused in part by increases of synaptic activity and NMDA-receptor-dependent calcium spikes in apical tuft dendrites. Furthermore, local inhibitory interneuron networks shifted their activity in favor of pyramidal neuron hyperactivity: somatostatin-expressing and parvalbumin-expressing inhibitory neurons reduced their activity, whereas vasoactive intestinal polypeptide-expressing interneurons increased their activity. Pharmacogenetic activation of somatostatin-expressing cells reduced pyramidal neuron hyperactivity and reversed mechanical allodynia. These findings reveal cortical circuit changes that arise during the development of neuropathic pain and identify the activation of specific cortical interneurons as therapeutic targets for chronic pain treatment.

It is well established that visual deprivation has a profound impact on the responsiveness of neurons in the developing visual cortex. The effect of visual deprivation on synaptic connectivity remains unclear. Using transcranial two-photon microscopy, we examined the effect of visual deprivation and subsequent recovery on dendritic spine remodeling of layer 5 pyramidal neurons in the mouse primary visual cortex. We found that monocular deprivation (MD), but not binocular deprivation (BD), increased dendritic spine elimination over 3 days in the binocular region of 4-week-old adolescent mice. This MD-induced dendritic spine elimination persisted during subsequent 2-4 days of binocular recovery. Furthermore, we found that average dendritic spine sizes were decreased and increased following 3-day MD and BD, respectively. These spine size changes induced by MD or BD tended to be reversed during subsequent binocular recovery. Taken together, these findings reveal differential effects of MD and BD on synaptic connectivity of layer 5 pyramidal neurons and underscore the persistent impact of MD on synapse loss in the developing visual cortex.

BACKGROUND: Microglia are known as resident immune cells in the brain. beta-amyloid (Abeta) plaques in the brain of Alzheimer's disease (AD) are surrounded by microglia, but whether and how microglia affect the formation and maintenance of plaques remains controversial. METHODS: We depleted microglia by injecting diphtheria toxin (DT) in CX 3 CR1 CreER/+ :R26 DTR/+ (CX 3 CR1-iDTR) mice crossed with APPswe/PSEN1dE9 (APP/PS1) mice. Intravital time-lapse imaging was performed to examine changes in the number and size of Congo Red-labeled amyloid plaques over 1-2 weeks. We also examined spine density and shaft diameter of dendrites passing through plaques in a PSAPP mouse model of AD (PS1 M146L line 6.2 x Tg2576) crossed with Thy1 YFP H-line mice. RESULTS: We found that DT administration to CX 3 CR1-iDTR mice efficiently ablated microglia within one week and that microglia repopulated in the second week after DT administration. Microglia depletion didn't affect the number of amyloid plaques, but led to ~13% increase in the size of Abeta plaques within one week. Moreover, microglia repopulation was associated with the stabilization of plaque size during the second week. In addition, we found dendritic spine loss and shaft atrophy in the distal parts of dendrites passing through plaques. CONCLUSION: Our results demonstrate the important role of microglia in limiting the growth of Abeta plaques and plaque-associated disruption of neuronal connection.

Frontotemporal dementia (FTD) is the second most common dementia before 65 years of age. Haploinsufficiency in the progranulin (GRN) gene accounts for 10% of all cases of familial FTD. GRN mutation carriers have an increased risk of autoimmune disorders, accompanied by elevated levels of tissue necrosis factor (TNF) alpha. We examined behavioral alterations related to obsessive-compulsive disorder (OCD) and the role of TNFalpha and related signaling pathways in FTD patients with GRN mutations and in mice lacking progranulin (PGRN). We found that patients and mice with GRN mutations displayed OCD and self-grooming (an OCD-like behavior in mice), respectively. Furthermore, medium spiny neurons in the nucleus accumbens, an area implicated in development of OCD, display hyperexcitability in PGRN knockout mice. Reducing levels of TNFalpha in PGRN knockout mice abolished excessive self-grooming and the associated hyperexcitability of medium spiny neurons of the nucleus accumbens. In the brain, PGRN is highly expressed in microglia, which are a major source of TNFalpha. We therefore deleted PGRN specifically in microglia and found that it was sufficient to induce excessive grooming. Importantly, excessive grooming in these mice was prevented by inactivating nuclear factor kappaB (NF-kappaB) in microglia/myeloid cells. Our findings suggest that PGRN deficiency leads to excessive NF-kappaB activation in microglia and elevated TNFalpha signaling, which in turn lead to hyperexcitability of medium spiny neurons and OCD-like behavior.

The functions and underlying mechanisms of rapid eye movement (REM) sleep remain unclear. Here we show that REM sleep prunes newly formed postsynaptic dendritic spines of layer 5 pyramidal neurons in the mouse motor cortex during development and motor learning. This REM sleep-dependent elimination of new spines facilitates subsequent spine formation during development and when a new motor task is learned, indicating a role for REM sleep in pruning to balance the number of new spines formed over time. Moreover, REM sleep also strengthens and maintains newly formed spines, which are critical for neuronal circuit development and behavioral improvement after learning. We further show that dendritic calcium spikes arising during REM sleep are important for pruning and strengthening new spines. Together, these findings indicate that REM sleep has multifaceted functions in brain development, learning and memory consolidation by selectively eliminating and maintaining newly formed synapses via dendritic calcium spike-dependent mechanisms.

Microglia and peripheral monocytes contribute to hypersensitivity in rodent models of neuropathic pain. However, the precise respective function of microglia and peripheral monocytes has not been investigated in these models. To address this question, here we combined transgenic mice and pharmacological tools to specifically and temporally control the depletion of microglia and monocytes in a mouse model of spinal nerve transection (SNT). We found that although microglia and monocytes are required during the initiation of mechanical allodynia or thermal hyperalgesia, these cells may not be as important for the maintenance of hypersensitivity. Moreover, we demonstrated that either resident microglia or peripheral monocytes are sufficient in gating neuropathic pain after SNT. We propose that resident microglia and peripheral monocytes act synergistically to initiate hypersensitivity and promote the transition from acute to chronic pain after peripheral nerve injury.

Chitooligosaccharide (COS) has been shown to induce fibroblast apoptosis, indicating that it could be used as a material to inhibit scar formation. In the present study, we used a mouse model of sciatic nerve injury (SNI) to determine the role of COS in scar inhibition and functional recovery. The animals were divided into three groups: SNI, SNI + vehicle, and SNI + COS group. We performed a series of functional and histological examinations at ctrl, 0 min, 14 days, and 42 days, including behavioral recovery, percentage of regenerating axons, degree of scar formation, vascular changes, type I and type III collagen ratio, and percentage of demyelinated axons. The SNI + COS group exhibited better recovery of sensory and motor function and less scar formation. Two-photon microscopy showed that the percentage of regenerating axons was highest in the SNI + COS group at 14 and 42 days. Our results suggested that COS can inhibit scar formation and enhance functional recovery by inducing fibroblast death, altering the proportion of different vascular diameters, changing the ratio of type I/type III collagen, and reducing the percentage of demyelinated axons. COS might be a useful drug in the treatment of SNI to reduce scar formation, but additional research is required to clarify the relevant molecular pathways.

Microglia, the principal resident immune cell of the CNS, exert significant influence on neurons during development and in pathological situations. However, if and how microglia contribute to normal neuronal function in the mature uninjured CNS is not well understood. We used the model of the adult mouse retina, a part of the CNS amenable to structural and functional analysis, to investigate the constitutive role of microglia by depleting microglia from the retina in a sustained manner using genetic methods. We discovered that microglia are not acutely required for the maintenance of adult retinal architecture, the survival of retinal neurons, or the laminar organization of their dendritic and axonal compartments. However, sustained microglial depletion results in the degeneration of photoreceptor synapses in the outer plexiform layer, leading to a progressive functional deterioration in retinal light responses. Our results demonstrate that microglia are constitutively required for the maintenance of synaptic structure in the adult retina and for synaptic transmission underlying normal visual function. Our findings on constitutive microglial function are relevant in understanding microglial contributions to pathology and in the consideration of therapeutic interventions that reduce or perturb constitutive microglial function. SIGNIFICANCE STATEMENT: Microglia, the principal resident immune cell population in the CNS, has been implicated in diseases in the brain and retina. However, how they contribute to the everyday function of the CNS is unclear. Using the model of the adult mouse retina, we examined the constitutive role of microglia by depleting microglia from the retina. We found that in the absence of microglia, retinal neurons did not undergo overt cell death or become structurally disorganized in their processes. However, connections between neurons called synapses begin to break down, leading to a decreased ability of the retina to transmit light responses. Our results indicate that retinal microglia contribute constitutively to the maintenance of synapses underlying healthy vision.