Bowdoin Science Journal

Hailey Ryan '26

Sex-Specific Brain Responses: How Chronic Stress Reshapes Astrocytes Differently in Males and Females

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Chronic stress is one of the major precursors to numerous neuropsychiatric disorders, such as depression. Women are twice as likely to be affected by mood disorders and respond differently than men to current available treatments. Nonetheless, many preclinical studies are conducted only in male rodents. Investigating the sex-specific responses to stress is critical to identifying mechanisms underlying mood disorders and moving towards developing treatments suitable for sex differences (Zhang et al. 2024). 

Chronic stress is associated with increased inflammation in the brain. The astrocyte is a type of cell in the nervous system that regulates inflammation in the brain. Astrocytes are important for keeping neurons alive, maintaining homeostasis, and secreting cytokines that regulate proinflammatory factors in the brain. The morphological changes of reactive astrocytes can tell us about the inflammation in the brain. These changes include the number of branches the astrocyte has; the more branches it has, the more reactive the astrocyte is, which is a sign of increased stress and inflammation in the brain. Chronic stress can lead to hyperactivation of astrocytes, impairing their ability to control and limit the spread of inflammation. Remodeling of astrocytes through changes in their cellular branching (more cellular branching) has been observed in suicide victims and preclinical chronic stress models. A recent study at Rowan University investigated sex-specific astrocyte responses to chronic stress in brain regions associated with mood disorders. 

Figure 1. Non-reactive vs. Reactive astrocytes. Reactive astrocytes have thicker cell bodies and processes. Astrocytes become reactive in response to injury, as well as to chronic stress. Adapted from: Pekny M, Wilhelmsson U, Pekna M. 2014. The dual role of astrocyte activation and reactive gliosis. Neuroscience Letters. 565:30–38. 

The study used the unpredictable chronic mild stress (UCMS) model to model chronic stress or a lipopolysaccharide (LPS) injection to model systemic inflammation. The UCMS model in rodents induces behavioral symptoms commonly associated with clinical depression as well as physiological and neurological changes that are associated with depression, such as hypertension and learned helplessness. The protocol involves randomized, daily exposures to different stressors, such as removal or bedding, social stresses, or predator sounds/smells. This model allows for an in-depth investigation of changes associated with chronic-stress induced depression (Frisbee et al. 2015). The LPS injection leads to neuroinflammation, sickness behavior, and cognitive impairment; it is a model often used to study neuroinflammation-associated diseases in mice. LPS causes inflammation because it activates microglia, which are the immune cells in the nervous system and play a large role in neuroinflammation (Zhao et al. 2019). 

Male and female mice were randomly assigned to the control group (saline), a 4 hour LPS injection (4LPS), a 24 LPS injection (24LPS), no stress (NS), or stress (UCMS). They measured GFAP (a biological marker of astrocyte reactivity) fluorescent intensity for astrocyte expression and quantified branch points to assess astrocyte complexity in different brain regions. 

The astrocyte complexity was investigated in the hippocampus, amygdala, and hypothalamus. The hippocampus and amygdala are critical brain regions in regulating physiological and behavioral stress processes, which can be useful in the short-term but detrimental in the long-term. In the short term, they help the body respond to stressors in order to maintain homeostasis. However, these stress mechanisms can lead to long-term dysregulation of this process as they promote maladaptive damage on the body and brain under chronically stressful conditions, which compromises resiliency and health. The amygdala is involved in detecting and responding to threats in the environment; the hippocampus is important for memory. These regions work together to make emotional and salient memories strong and long-lasting. Inflammation may hinder learning and memory through structural remodeling of the hippocampus (McEwen and Gianaros 2010). The hypothalamus, which is part of the hypothalamic-pituitary-adrenal (HPA) axis, is essential for mediating the stress response, primarily through the release of stress hormones (Bao et al. 2008). 

Figure 2.  The hypothalamus, hippocampus, and amygdala. Adapted from: https://www.brainframe-kids.com/emotions/facts-brain.htm.

The study found that chronic stress-induced morphological changes in astrocytes occurred in all brain regions that were looked at, and that the effects of chronic stress were both region and sex specific. Females had greater stress or inflammation-induced astrocyte activation in the hypothalamus, hippocampus, and amygdala than males. This indicates that chronic stress induces astrocyte activation that could drive sex-specific differences, which may contribute to the sex differences of mood disorders and disease. 

To better assess astrocyte reactivity, they used the ramification index, which is the ratio between the total number of primary branches and the branch maximum. It indicates how ramified the astrocytes are, as more branches (more ramified) indicates more reactive astrocytes. The ramification index indicated that astrocytes were significantly ramified due to chronic stress or LPS injection. They also conducted analysis of morphological changes, which provides the strongest evidence of astrocyte reactivity. The following results demonstrate the sex differences due to stress in branch point and terminal point morphology measurements. 

In the chronic-stress induced inflammatory environment (UCMS), there was higher astrocyte activation in the female hippocampus and hypothalamus, as demonstrated by increased branch points (Figure 3). 

Figure 3. UCMS and LPS activate astrocytes in the hypothalamus by inducing morphological changes. A. Representative images of female astrocytes in the hypothalamus in each condition (saline, no stress, 4 hours post-LPS, 24 hours post-LPS, and UCMS). D. Branching points in the hypothalamus. There were significant differences between treatment and between sex. There were significantly more branch points for females in the UCMS condition than for males. Adapted from: Zhang AY, Elias E, Manners MT. 2024. Sex-dependent astrocyte reactivity: Unveiling chronic stress-induced morphological changes across multiple brain regions. Neurobiology of Disease. 200:106610.

In the amygdala and hippocampus in the 24LPS condition, there was increased astrocyte reactivity in females compared to males. This indicates that females are more susceptible to chronic systemic inflammation than males in these brain regions (Figure 4). 

Figure 4. UCMS and LPS activate astrocytes in the amygdala by inducing morphological changes. A. Representative images of female astrocytes in the amygdala in the saline, no stress, 4LPS, 24LPS, and UCMS groups. D. Analysis of branch points in the amygdala. There were significant differences between treatment and between sex. Females had significantly more branching points in the 24LPS condition than males. Adapted from: Zhang AY, Elias E, Manners MT. 2024. Sex-dependent astrocyte reactivity: Unveiling chronic stress-induced morphological changes across multiple brain regions. Neurobiology of Disease. 200:106610.

Astrocytes work in tandem with other cells in the nervous system, including microglia, to regulate various processes. Microglia, the cells in the nervous system important for immune response, are also important in the inflammatory response in the brain. Reactive microglia can activate astrocytes by secreting cytokines. Blocking microglia may be able to decrease the number of reactive astrocytes and apply a protective effect against inflammation in the brain. Future studies can look at the activation of microglia and how microglia and astrocytes interact in the chronic stress model. 

Overall, it is important that this study looked at sex differences since there is such a disparity among mental health disorders and treatment in women and men. Understanding the mechanisms behind the sex differences can improve the development of new medications for stress-related disorders so that both men and women can be correctly treated. 

Moreover, female susceptibility to chronic stress may mediate the increased risk for Alzheimer’s Disease. The different biochemical responses to stress, such as activity of the hypothalamic-pituitary-adrenal (HPA) axis and female-biased increases in molecules associated with AD, between females and males could be a sex-dependent risk factor for AD. Female-specific alterations in inflammation and microglial function are proposed to be one reason, but this needs more investigation (Yan et al. 2018). Understanding sex-specific disease mechanisms is essential for the development of personalized medicine, which is the use of an individual’s genetic profile to prevent, diagnose, and treat disease. Differences in mechanisms of disease between sexes will likely require different drugs for men and women to treat a variety of psychiatric and neurological disorders (Bangasser and Wicks 2017). 

 

References. 

Bangasser DA, Wicks B. 2017. Sex-specific mechanisms for responding to stress. Journal of Neuroscience Research. 95(1–2):75–82. 

Bao A-M, Meynen G, Swaab DF. 2008. The stress system in depression and neurodegeneration: Focus on the human hypothalamus. Brain Research Reviews. 57(2):531–553. 

Emotion Facts: Emotions in the Brain. [accessed 2024 Oct 29]. https://www.brainframe-kids.com/emotions/facts-brain.htm.

Frisbee JC, Brooks SD, Stanley SC, d’Audiffret AC. 2015. An Unpredictable Chronic Mild Stress Protocol for Instigating Depressive Symptoms, Behavioral Changes and Negative Health Outcomes in Rodents. J Vis Exp.(106):53109. 

McEwen BS, Gianaros PJ. 2010. Central role of the brain in stress and adaptation: Links to socioeconomic status, health, and disease. Annals of the New York Academy of Sciences. 1186:190. 

Pekny M, Wilhelmsson U, Pekna M. 2014. The dual role of astrocyte activation and reactive gliosis. Neuroscience Letters. 565:30–38. 

Tynan RJ, Naicker S, Hinwood M, Nalivaiko E, Buller KM, Pow DV, Day TA, Walker FR. 2010. Chronic stress alters the density and morphology of microglia in a subset of stress-responsive brain regions. Brain, Behavior, and Immunity. 24(7):1058–1068. 

Yan Y, Dominguez S, Fisher DW, Dong H. 2018. Sex differences in chronic stress responses and Alzheimer’s disease. Neurobiology of Stress. 8:120–126. 

Zhang AY, Elias E, Manners MT. 2024. Sex-dependent astrocyte reactivity: Unveiling chronic stress-induced morphological changes across multiple brain regions. Neurobiology of Disease. 200:106610. 

Zhao J, Bi W, Xiao S, Lan X, Cheng X, Zhang J, Lu D, Wei W, Wang Y, Li H, et al. 2019. Neuroinflammation induced by lipopolysaccharide causes cognitive impairment in mice. Sci Rep. 9(1):5790. 

Saffron Can Reduce Cognitive Impairment and Biomarkers of Alzheimer’s Disease

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Saffron is a popular food condiment known to have a positive effect against neurodegenerative diseases such as dementia, Alzheimer’s, and Parkinson’s. Saffron could potentially alleviate some of the biological hallmarks of Alzheimer’s disease, including the accumulation of Aβ plaques outside of cells and neurofibrillary tangles (NFTs) inside of cells. It could also target the neurotransmitter system that involves acetylcholine (ACh), which when disrupted can lead to the progression of dementia. This system, called the cholinergic system, is essential for attention, learning, and memory. ACh plays a role in memory acquisition, encoding, and consolidation (Patel et al. 2024). 

A recent study looked at how saffron extract (CSE) supplementation affects behavioral activity, memory, Aβ plaques, and NFTs in rats. The study also investigated how saffron affects a disrupted cholinergic system. When the system breaks down, there is an increase in Aβ plaques and NFTs. There is also an increase in AChE, which is the protein that breaks down ACh. All of these factors can ultimately contribute to Alzheimer’s. The current study investigated how CSE could reduce these effects and ultimately prevent progression of dementia-related diseases (Patel et al. 2024).  

In order to study the effects of CSE, the researchers used the drug scopolamine to disrupt the cholinergic system in rats. Scopolamine mimics the memory loss observed in dementia, and it is often used to study the cholinergic system in animal models of dementia. It primarily impairs the acquisition and encoding of memories (Paul 2019). Scopolamine increases the level of AChE in the brain, which means that more ACh is broken down. With less ACh available, the cholinergic system is disrupted, causing cognitive impairments (Figure 1). The study observed the potential of CSE to reduce this disturbance. They used behavioral tests to observe memory and spatial acquisition. Blood samples and brain samples were collected for examination (Patel et al. 2024). 

Figure 1. The role of acetylcholinesterase (AChE). AChE breaks down acetylcholine (ACh) as it moves from one neuron to another. An increase in AChE leads to a decrease in ACh. (Adapted from What is the role of acetylcholinesterase at a synapse?)

The researchers found that CSE supplementation significantly improved behavioral activity and memory in rats. CSE improved performance on the reversal task, which requires the ability to switch between learned and new responses, a reflection of cognitive flexibility. 

Importantly, the study offers insight into the biological parameters that lead to the improved cognitive abilities. CSE lowered the levels of AChE, meaning that less acetylcholine was broken down. The dose of 20 mg/kg of CSE lowered the amount of AChE in the scopolamine-treated rats as significantly as Rivastigmine did (Figure 2). Rivastigmine is a pharmaceutical drug currently used to treat symptoms of Alzheimer’s by inhibiting AChE (Emre et al. 2010). An increase in the level of acetylcholine can improve memory, as demonstrated by the improved performance of the rats on the memory tests. ACh has been shown to promote nerve conduction related to memory and learning ability, helping memories get encoded and consolidated in the brain. 

Figure 2. CSE supplementation lowered the levels of AChE and inhibited its effects in rats who received scopolamine administration. Thus, less acetylcholine was broken down and disruption of the cholinergic system was attenuated. This can lead to an improvement in memory (Adapted from Patel et al, 2024). 

CSE supplementation also significantly reduced the formation of Aβ plaques and NFTs, the two classical pathological hallmarks of Alzheimer’s in the hippocampus in the brain. The 20 mg/kg dose of CSE produced the most significant reduction. CSE reduced the number of Aβ plaques and NFTs more than Rivastigmine did. In fact, the Rivastigmine group still had significant levels of NFTs and Aβ plaques (Figure 3). Accumulation of Aβ plaques and NFTs lead to neuronal dysfunction and ultimately dementia. Moreover, AChE and Aβ can bind together to form complexes that exaggerate neurodegeneration even more. Thus, if CSE can attenuate their formation, neuronal function may be preserved, preventing the progression of dementia.  

Figure 3. CSE supplementation lowered the density of Aβ plaques in rats who received scopolamine administration. This can lead to an improvement in memory (Adapted from Patel et al, 2024). 

Figure 4. Summary of the article. Scopolamine administration led to a disruption of the cholinergic system (a part of the nervous system involving acetylcholine that helps with memory and learning), neuroinflammation, and accumulation of NFT and Aβ plaques. When CSE (saffron extract) was administered, the rats’ memory improved and the biomarkers of Alzheimer’s were reduced (Adapted from Patel et al, 2024). 

Because CSE supplementation reduces the formation of Aβ plaque and NFTs in the hippocampus, inhibits the activity of the enzyme that breaks down acetylcholine, and counters neuroinflammation, a saffron-based botanical supplement could be used to help manage dementia. Currently, manufactured drugs such as donepezil and memantine have been used to treat symptoms of Alzheimer’s, but saffron could potentially offer a safer alternative that is just as effective (D’Onofrio et al. 2021). The current study tested the effects of CSE against Rivastigmine, another drug that is approved to treat symptoms of Alzheimer’s. CSE produced the same improvements as Rivastigmine. Rivastigmine, along with other chemical drugs that inhibit AChE in order to treat Alzheimer’s, has been shown to cause gastrointestinal side effects and does not have great patient compliance (Emre et al. 2010). Saffron supplementation could offer an effective treatment without such side effects, increasing quality of life. The research offers promise that saffron can improve the cognitive impairments associated with Alzheimer’s and dementia. 

 

Literature Cited

Emre M, Bernabei R, Blesa R, Bullock R, Cunha L, Daniëls H, Dziadulewicz E, Förstl H, Frölich L, Gabryelewicz T, et al. 2010. Drug Profile: Transdermal Rivastigmine Patch in the Treatment of Alzheimer Disease. CNS Neurosci Ther. 16(4):246–253. 

D’Onofrio G, Nabavi SM, Sancarlo D, Greco A, Pieretti S. 2021. Crocus Sativus L. (Saffron) in Alzheimer’s Disease Treatment: Bioactive Effects on Cognitive Impairment. Curr Neuropharmacol. 19(9):1606–1616. 

Patel KS, Dharamsi A, Priya M, Jain S, Mandal V, Girme A, Modi SJ, Hingorani L. 2024a. Saffron (Crocus sativus L.) extract attenuates chronic scopolamine-induced cognitive impairment, amyloid beta, and neurofibrillary tangles accumulation in rats. Journal of Ethnopharmacology. 326:117898. 

Paul J. 2019. Chapter 5 – Experimental Medicine Approaches in CNS Drug Development. In: Nomikos GG, Feltner DE, editors. Handbook of Behavioral Neuroscience. Vol. 29. Elsevier. (Translational Medicine in CNS Drug Development). p. 63–80. [accessed 2024 Feb 25]. 

What is the role of acetylcholinesterase at a synapse? | Socratic. Socratic.org. [accessed 2024 Mar 21]. 

Retrieving Forgotten Memories With Optogenetics

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Alzheimer’s disease is a neurodegenerative disorder in which patients experience a progressive decline in memory (Roy et. al, 2016). The early stages involve struggles to remember mostly episodic memories, which are memories of personal experiences that are linked to activity in the hippocampus. To discover potential treatments, scientists have tried to better understand the biological mechanisms behind the disease. Optogenetic activation of engram cells and calculation of dendritic spine density are two methods to research the disease. Dendritic spines are extensions from dendrites, which are the structures that receive signals from other neurons and pass them to the cell body. Memory is associated with changes in dendritic spines or the formation of new ones, which makes scientists believe that they serve as sites for memory formation and storage (Roy et. al, 2016). Engram cells are biological traces of established memories that store the information and are reactivated when memories are retrieved (Ortega-de San Luis et al., 2022). Optogenetics is a technique in which genes for light sensitive proteins are injected into specific neurons in the brain so that they can be activated by laser lights that open ion channels (Figure 1). 

Figure 1. The process of optogenetics (Adapted from Buchen, 2010). 

A 2016 study affiliated with the Massachusetts Institute of Technology used optogenetics to activate engram cells in the hippocampus of mouse models of early Alzheimer’s disease (AD) (Roy et al., 2016). With direct activation of hippocampal cells, the mice were able to retrieve the memories they had forgotten, indicating that the issue is retrieving memories; the memories are still available in the brain, but there is a problem in recalling them. Amnesia in early AD mice is also correlated with a reduction of spine density of engram cells in the hippocampus. The study showed that restoring dendritic spine density with optogenetic activation allows for the retrieval of long-term memory. Spine density was restored as the optogenetic activation allowed sodium ions to flow into the neurons and consequently depolarized them (meaning the cell is less negatively charged). This depolarization triggers long term potentiation, which is the process of strengthening connections between neurons that underlies memory and learning. Thus, restoration of spine density could lead to an effective strategy for treating memory loss in patients with early Alzheimer’s disease. 

In the experiment, the scientists labeled engram cells in the mouse models of Early Alzheimer’s disease and tested memory recall with contextual fear conditioning and long-term memory testing. Contextual fear conditioning is when mice are trained to associate a particular context (a cage) with a shock, creating a memory of the association in the hippocampus. Long-term memory was tested by placing them back into the cage and measuring their freezing behavior, a commonly used measure of fear. They also calculated dendritic spine density. 

The researchers found that optogenetic activation of memory engrams restores fear memory in early AD mice and that reversal of engram-specific spine deficits rescues memory in early AD mice. The diagram below shows the process of fear conditioning (panel t): the mice were trained to associate a shock with a certain context, and then their long term memory was tested by placing them back into the cage and observing their “freeze response,” which demonstrates that they feel fear in the cage because of the memory of the shock in the cage. The graphs show that when the engrams were activated optogenetically, the mice froze more than they did when the engrams were not activated, indicating that the optogenetic activation of the engram cells restored the mice’s memory of the fearful context. 

Figure 2. Optogenetic activation of memory engrams restores fear memory in early AD mice (Adapted from Roy et al., 2016, Fig. 1). 

The results of this study provide directions and hope for future treatments for Alzheimer’s Disease. If the amnesia is due to retrieval impairments, memory could be restored by technologies involving brain stimulation, like the optogenetic activation did in the mouse models. However, optogenetics is currently a technique only possible in animals since it is invasive, so further research will have to be done to discover a technique employing the principle in a more plausible way for humans, perhaps by finding a way to restore spine density in engram cells. There are other limitations when it comes to future treatments for Alzheimer’s because of the fact that they studied only early AD mice and episodic memory. Even though they showed that amnesia in early AD mice impairs memory retrieval, long-term memory storage in advanced stages of AD may also be impaired and eventually lost as neuronal degeneration progresses. Also, humans with early AD often exhibit non-episodic memory deficits as well, which involve brain structures outside of the medial temporal lobe that the current study did not investigate. However, overall, the findings contribute to a better understanding of memory retrieval deficits in early cases of AD, which may also apply to other neurological diseases in which patients have difficulty with memory retrieval, such as Huntington’s Disease. 

 

Literature Cited

Buchen, L. (2010). Neuroscience: Illuminating the brain. Nature, 465(7294), Article 7294. https://doi.org/10.1038/465026a

Ortega-de San Luis, C., & Ryan, T. J. (2022b). Understanding the physical basis of memory: Molecular mechanisms of the engram. The Journal of Biological Chemistry, 298(5), 101866. https://doi.org/10.1016/j.jbc.2022.101866

Roy, D. S., Arons, A., Mitchell, T. I., Pignatelli, M., Ryan, T. J., & Tonegawa, S. (2016d). Memory retrieval by activating engram cells in mouse models of early Alzheimer’s disease. Nature, 531(7595), 508–512. https://doi.org/10.1038/nature17172

Yuhas, D. (n.d.). Forgotten Memories May Remain Intact in the Brain. Scientific American. Retrieved November 4, 2023, from https://www.scientificamerican.com/article/forgotten-memories-may-remain-intact-in-the-brain/