Researchers Identify Potential Treatment for Patients at Risk for Alzheimer’s Disease
Neurolucida 360 Used to Analyze Dendrites and Dendritic Spines
Amyloid plaques and tau tangles are the hallmarks of Alzheimer’s disease (AD) pathology, but synapse loss is what causes cognitive decline, scientists say. In a paper published in Science Signaling, researchers at the Herskowitz Lab, at the University of Alabama at Birmingham, used Neurolucida 360 to analyze spine density and dendritic length in hAPP mice — a mouse model of AD. Their findings describe a treatment that could protect against synapse loss and prevent the onset of dementia in patients at risk for Alzheimer’s disease.
Targeting LIMK1 to Protect Against Dendritic Damage
In their study, the scientists targeted LIMK1, an enzyme that regulates the size and density of dendritic spines. Previous studies have shown that in animal models of AD, LIMK1 activity is increased, causing synaptic hyperactivity and dendritic damage. After confirming this phenomenon, the research team set out to find a way to inhibit LIMK1, which lies downstream of two other important players in dementia pathology — the Rho-associated kinases known as ROCK1 and ROCK2.
Representative maximum-intensity high-resolution confocal microscope images of dye-filled dendrites, from CA1 hippocampal neurons in mice, after deconvolution and corresponding 3D digital reconstruction models of dendrites. Scale bar, 5 μm. Colors in digital reconstructions correspond to dendritic protrusion classes: blue, thin spines; orange, stubby spines; green, mushroom spines; and yellow, dendritic filopodia.
Previous studies have shown that severe side effects including fatally low blood pressure are associated with the inhibition of ROCK1 and ROCK2, so the researchers looked further down the signaling pathway to the LIMK1 point, potentially discovering a truly valid target in the fight to prevent dementia onset.
Since LIMK1 has also been a target in cancer treatment, the researchers turned to SR7826, an experimental drug currently in development to treat cancer patients. They found that administering SR7826 suppressed LIMK1 activity and protected dendritic morphology against the damage commonly seen in a brain afflicted with dementia. By reconstructing the mouse neurons with Neurolucida 360, they observed increased dendritic spine length and density in the experimental group, compared to controls.
Using Neurolucida 360 to Analyze Dendritic Spine Morphology
Herskowitz and his team have used Neurolucida 360 in previous studies, and used the software extensively throughout the current study to image and quantify dendritic morphology.
“Neurolucida 360 is a remarkable system that has provided us with tools to study dendritic architecture in cultured neurons, rodent models, and humans, with outstanding precision and detail,” said principal investigator Jeremy Herskowitz, Ph.D.
Their first step was to confirm that increased ROCK1 or ROCK2 activity caused detrimental structural effects on dendritic spines. To do this, they analyzed rat hippocampal neurons that had been transfected with either green fluorescent protein (GFP), an actin-binding peptide, ROCK1, or ROCK2. Controls were transfected with empty vectors.
Using Neurolucida 360, they digitally generated neuron reconstructions and quantified dendritic spine length and density in all of the experimental groups. Neurons expressing ROCK1 showed significantly reduced spine length compared to vector or GFP controls, while neurons expressing ROCK2 showed significantly reduced spine density.
They then used Neurolucida 360 to determine that two different mechanisms dictate the distinct impacts ROCK1 and ROCK2 have on dendritic structure — ROCK1 kinase activity regulates spine length through myosin-actin pathways, whereas ROCK2 kinase activity controls spine density through LIMK1-cofilin-actin signaling.
Further experiments used Neurolucida 360 to analyze dendrites of neurons injected with both amyloid-β oligomers and a virus expressing ROCK1- or ROCK2-targeted RNA. This allowed the scientists to discern that ROCK2, and not ROCK1, works with Aß oligomers to induce spine degeneration.
“Treatment of hAPP mice with a LIMK1 inhibitor rescued Aβ-induced hippocampal spine loss and morphologic aberrations. Our data suggest that therapeutically targeting LIMK1 may provide dendritic spine resilience to Aβ and therefore may benefit cognitively normal patients that are at high risk for developing dementia. (2019, Henderson et al)
Henderson, BW., Greathouse, KM., Ramdas, R., Walker, CK., Rao, TC., Bach, SV., Curtis, KA., Day, JJ., Mattheyses, AL., Herskowitz, JH. 2019. Pharmacologic inhibition of LIMK1 provides dendritic spine resilience against β-amyloid. Science Signaling Vol. 12, Issue 587 DOI: 10.1126/scisignal.aaw9318 (https://stke.sciencemag.org/content/12/587/eaaw9318)