Delivering progranulin to neuronal lysosomes protects against excitotoxicity.

Loss-of-function mutations in progranulin (GRN) are a major genetic cause of frontotemporal dementia (FTD), possibly due to loss of progranulin's neurotrophic and anti-inflammatory effects. Progranulin promotes neuronal growth and protects against excitotoxicity and other forms of injury. It is unclear if these neurotrophic effects are mediated through cellular signaling or through promotion of lysosomal function. Progranulin is a secreted proprotein that may activate neurotrophic signaling through cell-surface receptors. However, progranulin is efficiently trafficked to lysosomes and is necessary for maintaining lysosomal function. To determine which of these mechanisms mediates progranulin's protection against excitotoxicity, we generated lentiviral vectors expressing progranulin (PGRN) or lysosome-targeted progranulin (L-PGRN). L-PGRN was generated by fusing the LAMP-1 transmembrane and cytosolic domains to the C-terminus of progranulin. L-PGRN exhibited no detectable secretion, but was delivered to lysosomes and processed into granulins. PGRN and L-PGRN protected against NMDA excitotoxicity in rat primary cortical neurons, but L-PGRN had more consistent protective effects than PGRN. L-PGRN's protective effects were likely mediated through the autophagy-lysosomal pathway. In control neurons, an excitotoxic dose of NMDA stimulated autophagy, and inhibiting autophagy with 3-methyladenine reduced excitotoxic cell death. L-PGRN blunted the autophagic response to NMDA and occluded the protective effect of 3-methyladenine. This was not due to a general impairment of autophagy, as L-PGRN increased basal autophagy and did not alter autophagy after nutrient starvation. These data show that progranulin's protection against excitotoxicity does not require extracellular progranulin, but is mediated through lysosomes, providing a mechanistic link between progranulin's lysosomal and neurotrophic effects.

Race and the Ivory Tower: An Antiracism Exercise for an Undergraduate Neuroscience Classroom.

This article details an antiracism exercise completed in an introductory undergraduate neuroscience class. Students completed an online pre-class multimedia module entitled "Race and the Ivory Tower" covering racism in science and medicine, the neuroscience behind bias, and the impact of race and racism on health outcomes. The module included two videos, one podcast, and a peer-reviewed journal article, alongside several optional additional resources written for both academic and lay audiences. After completing the module, students participated in an open-ended online discussion followed by an anonymous survey to elicit feedback on the exercise. As a continuation of the antiracism exercise, students researched and reported on the work of a Black or nonwhite Hispanic/Latino scientist for a final project later in the semester. Sixty-eight of 69 students participated in the discussion, and the majority discussed the neuroscience of bias and public health effects of racism. Most students also discussed the importance of the module contents or further questions that they would explore. Sixty of 69 students answered the anonymous survey, where most students reported a better understanding of racism after interacting with the content. Additionally, most students felt better prepared to discuss racism in science and medicine and more able to identify unconscious bias. Finally, students reported that they enjoyed the module contents and online discussion. Overall, this exercise effectively introduced students to the ongoing challenge of racism in science and medicine through both scientific and sociological lenses. Students recognized the collective importance of the content, which was our goal as they represent the future leaders in neuroscience and medicine and should be equipped to address leading issues within their field.

A peptide inhibitor of Tau-SH3 interactions ameliorates amyloid-ß toxicity.

The microtubule-associated protein Tau is strongly implicated in Alzheimer's disease (AD) and aggregates into neurofibrillary tangles in AD. Genetic reduction of Tau is protective in several animal models of AD and cell culture models of amyloid-ß (Aß) toxicity, making it an exciting therapeutic target for treating AD. A variety of evidence indicates that Tau's interactions with Fyn kinase and other SH3 domain-containing proteins, which bind to PxxP motifs in Tau's proline-rich domain, may contribute to AD deficits and Aß toxicity. Thus, we sought to determine if inhibiting Tau-SH3 interactions ameliorates Aß toxicity. We developed a peptide inhibitor of Tau-SH3 interactions and a proximity ligation assay (PLA)-based target engagement assay. Then, we used membrane trafficking and neurite degeneration assays to determine if inhibiting Tau-SH3 interactions ameliorated Aß oligomer (Aßo)-induced toxicity in primary hippocampal neurons from rats. We verified that Tau reduction ameliorated Aßo toxicity in neurons. Using PLA, we identified a peptide inhibitor that reduced Tau-SH3 interactions in HEK-293 cells and primary neurons. This peptide reduced Tau phosphorylation by Fyn without affecting Fyn's kinase activity state. In primary neurons, endogenous Tau-Fyn interaction was present primarily in neurites and was reduced by the peptide inhibitor, from which we inferred target engagement. Reducing Tau-SH3 interactions in neurons ameliorated Aßo toxicity by multiple outcome measures, namely Aßo-induced membrane trafficking abnormalities and neurite degeneration. Our results indicate that Tau-SH3 interactions are critical for Aßo toxicity and that inhibiting them is a promising therapeutic target for AD.

Development of small-molecule Tau-SH3 interaction inhibitors that prevent amyloid-ß toxicity and network hyperexcitability.

Alzheimer's disease (AD) is the leading cause of dementia and lacks highly effective treatments. Tau-based therapies hold promise. Tau reduction prevents amyloid-ß-induced dysfunction in preclinical models of AD and also prevents amyloid-ß-independent dysfunction in diverse disease models, especially those with network hyperexcitability, suggesting that strategies exploiting the mechanisms underlying Tau reduction may extend beyond AD. Tau binds several SH3 domain-containing proteins implicated in AD via its central proline-rich domain. We previously used a peptide inhibitor to demonstrate that blocking Tau interactions with SH3 domain-containing proteins ameliorates amyloid-ß-induced dysfunction. Here, we identify a top hit from high-throughput screening for small molecules that inhibit Tau-FynSH3 interactions and describe its optimization with medicinal chemistry. The resulting lead compound is a potent cell-permeable Tau-SH3 interaction inhibitor that binds Tau and prevents amyloid-ß-induced dysfunction, including network hyperexcitability. These data support the potential of using small molecule Tau-SH3 interaction inhibitors as a novel therapeutic approach to AD.

Circadian-mediated regulation of cardiometabolic disorders and aging with time-restricted feeding.

Circadian rhythms are present throughout biology, from the molecular level to complex behaviors such as eating and sleeping. They are driven by molecular clocks within cells, and different tissues can have unique rhythms. Circadian disruption can trigger obesity and other common metabolic disorders such as aging, diabetes, and cardiovascular disease, and circadian genes control metabolism. At an organismal level, feeding and fasting rhythms are key drivers of circadian rhythms. This underscores the bidirectional relationship between metabolism and circadian rhythms, and many metabolic disorders have circadian disruption or misalignment. Therefore, studying circadian rhythms may offer new avenues for understanding the etiology and management of obesity. This review describes how circadian rhythm dysregulation is linked with cardiometabolic disorders and how the lifestyle intervention of time-restricted feeding (TRF) regulates them. TRF reinforces feeding-fasting rhythms without reducing caloric intake and ameliorates metabolic disorders such as obesity and associated cardiac dysfunction, along with reducing inflammation. TRF optimizes the expression of genes and pathways related to normal metabolic function, linking metabolism with TRF's benefits and demonstrating the molecular link between metabolic disorders and circadian rhythms. Thus, TRF has tremendous therapeutic potential that could be easily adopted to reduce obesity-linked dysfunction and cardiometabolic disorders.

Rapamycin reduces neuronal mutant huntingtin aggregation and ameliorates locomotor performance in Drosophila.

Huntington's disease (HD) is a neurodegenerative disease characterized by movement and cognitive dysfunction. HD is caused by a CAG expansion in exon 1 of the HTT gene that leads to a polyglutamine (PQ) repeat in the huntingtin protein, which aggregates in the brain and periphery. Previously, we used Drosophila models to determine that Htt-PQ aggregation in the heart causes shortened lifespan and cardiac dysfunction that is ameliorated by promoting chaperonin function or reducing oxidative stress. Here, we further study the role of neuronal mutant huntingtin and how it affects peripheral function. We overexpressed normal (Htt-PQ25) or expanded mutant (Htt-PQ72) exon 1 of huntingtin in Drosophila neurons and found that mutant huntingtin caused age-dependent Htt-PQ aggregation in the brain and could cause a loss of synapsin. To determine if this neuronal dysfunction led to peripheral dysfunction, we performed a negative geotaxis assay to measure locomotor performance and found that neuronal mutant huntingtin caused an age-dependent decrease in locomotor performance. Next, we found that rapamycin reduced Htt-PQ aggregation in the brain. These results demonstrate the role of neuronal Htt-PQ in dysfunction in models of HD, suggest that brain-periphery crosstalk could be important to the pathogenesis of HD, and show that rapamycin reduces mutant huntingtin aggregation in the brain.

Alzheimer's disease risk gene BIN1 induces Tau-dependent network hyperexcitability.

Genome-wide association studies identified the BIN1 locus as a leading modulator of genetic risk in Alzheimer's disease (AD). One limitation in understanding BIN1's contribution to AD is its unknown function in the brain. AD-associated BIN1 variants are generally noncoding and likely change expression. Here, we determined the effects of increasing expression of the major neuronal isoform of human BIN1 in cultured rat hippocampal neurons. Higher BIN1 induced network hyperexcitability on multielectrode arrays, increased frequency of synaptic transmission, and elevated calcium transients, indicating that increasing BIN1 drives greater neuronal activity. In exploring the mechanism of these effects on neuronal physiology, we found that BIN1 interacted with L-type voltage-gated calcium channels (LVGCCs) and that BIN1-LVGCC interactions were modulated by Tau in rat hippocampal neurons and mouse brain. Finally, Tau reduction prevented BIN1-induced network hyperexcitability. These data shed light on BIN1's neuronal function and suggest that it may contribute to Tau-dependent hyperexcitability in AD.

Impaired ß-glucocerebrosidase activity and processing in frontotemporal dementia due to progranulin mutations.

Loss-of-function mutations in progranulin (GRN) are a major autosomal dominant cause of frontotemporal dementia. Most pathogenic GRN mutations result in progranulin haploinsufficiency, which is thought to cause frontotemporal dementia in GRN mutation carriers. Progranulin haploinsufficiency may drive frontotemporal dementia pathogenesis by disrupting lysosomal function, as patients with GRN mutations on both alleles develop the lysosomal storage disorder neuronal ceroid lipofuscinosis, and frontotemporal dementia patients with GRN mutations (FTD-GRN) also accumulate lipofuscin. The specific lysosomal deficits caused by progranulin insufficiency remain unclear, but emerging data indicate that progranulin insufficiency may impair lysosomal sphingolipid-metabolizing enzymes. We investigated the effects of progranulin insufficiency on sphingolipid-metabolizing enzymes in the inferior frontal gyrus of FTD-GRN patients using fluorogenic activity assays, biochemical profiling of enzyme levels and posttranslational modifications, and quantitative neuropathology. Of the enzymes studied, only ß-glucocerebrosidase exhibited impairment in FTD-GRN patients. Brains from FTD-GRN patients had lower activity than controls, which was associated with lower levels of mature ß-glucocerebrosidase protein and accumulation of insoluble, incompletely glycosylated ß-glucocerebrosidase. Immunostaining revealed loss of neuronal ß-glucocerebrosidase in FTD-GRN patients. To investigate the effects of progranulin insufficiency on ß-glucocerebrosidase outside of the context of neurodegeneration, we investigated ß-glucocerebrosidase activity in progranulin-insufficient mice. Brains from Grn-/- mice had lower ß-glucocerebrosidase activity than wild-type littermates, which was corrected by AAV-progranulin gene therapy. These data show that progranulin insufficiency impairs ß-glucocerebrosidase activity in the brain. This effect is strongest in neurons and may be caused by impaired ß-glucocerebrosidase processing.