Bishop, B, Morgan, E, Vargas, M, Ewing, S and Donley, DW

Department of Biology, Harding University, Searcy, AR

Introduction/Background. Microglia are highly plastic immune cells in the central nervous system that have multi-faceted responses to damage-associated signals. Chronic microglial activation and subsequent inflammation is linked to the progression of neurological diseases such as Alzheimer’s disease (AD) by potentiating rather suppressing damage. Despite a well characterized phenotype of microglia, mechanism(s) that control their dynamic responses in disease are still poorly understood.

Hypothesis/Goal of Study. To advance our understanding of the mechanisms underlying microglial activation in disease, we stimulate cultured microglia with iron and amyloid-beta due to their correlation with pathological inflammation and neurological disease progression. Amyloid-beta and iron accumulate concurrent with AD progression and are associated with detrimental inflammatory responses from microglia. However, the mechanism of how amyloid-beta and iron converge to induce microglial dysfunction remains elusive.

Methods and Results. The Normal Mucosa of Esophagus-Specific gene 1 (NMES1) protein and its gene, C15orf48, appear to be an intersection point of iron and amyloid-beta. Intriguingly, a proteomics analysis showed amyloid-beta stimulation increased expression of NMES1 while the addition of iron suppressed this effect in microglia. Further, we found that suppression of NMES1 leads to hyperactivation of microglia stimulated with amyloid-beta, suggesting that iron suppresses the ability of microglia to regulate inflammation downstream of amyloid-beta. These results indicate that NMES1 may be a critical regulator of inflammation in AD but more work is needed to elucidate how iron disrupts C15orf48/NMES1 function during disease.

Discussion/Conclusions. Our data expands on the emerging understanding of the impact of iron dysregulation on the inflammatory response of microglia to disease stimuli such as amyloid-beta. Broadly our work contributes to a better understanding of mechanisms underlying chronic microglial inflammation in the context of AD, and other neurological diseases.

Citation/Acknowledgements. This project was supported by the Harding University Department of Biology and by the Arkansas INBRE program, with a grant from the National Institute of General Medical Sciences, (NIGMS), P20 GM103429 from the National Institutes of Health.A.3: Cardiovascular Disease & Stroke.

José A. Lasalde-Dominicci1,2, Rolando Irizarry-Álvarez1, Randy Irizarry-Álvarez1, Emilio A. Cepeda-Terrasa1, Alanis Z. Pérez-Montalvo1, Manuel Delgado-Vélez1,2, Negin Martin3, and Jerrel L. Yakel4
1Department of Biology, University of Puerto Rico, San Juan, PR, 2Molecular Sciences Research Center, University of Puerto Rico, San Juan, PR, 3National
Institute of Environmental Health Sciences, National Institutes of Health, Bethesda, MD, 4Neurobiology Laboratory, National Institutes of Health, Bethesda, MD

Introduction/Background. Viral infections caused by HIV-1 and SARS-CoV-2 lead to the release of viral proteins within patients' bodies, initiating acute and chronic inflammatory processes that contribute to the emergence of comorbidities, ultimately diminishing patients' life expectancy. The cholinergic anti-inflammatory response (CAR) represents a vital neuroimmune mechanism hinging upon the expression of the neural receptor alpha7 nicotinic acetylcholine receptor (α7-nAChR) on macrophages.

Hypothesis/Goal of Study. This project seeks to ascertain whether the viral glycoproteins, gp120 (from HIV) and spike protein (from SARS-CoV-2), induce modifications in the expression of α7-nAChR within macrophages. Our hypothesis postulates that these viral proteins disrupt α7-nAChR expression in macrophages, potentially resulting in proinflammatory consequences by impeding CAP functionality.

Methods and Results. Monocyte-derived macrophages (MDMs) were sourced from healthy individuals and subsequently exposed to pathophysiologically relevant concentrations of the aforementioned viral glycoproteins. Subsequently, confocal imaging, following fluorescent labeling, was utilized to assess the expression of α7-nAChR. The outcomes of this investigation revealed that gp120 from HIV-1 prompted an elevation in α7-nAChR expression, while the spike protein from SARS-CoV-2 led to a reduction in α7-nAChR levels within macrophages.

Discussion/Conclusions. The observed disturbance in α7-nAChR levels among macrophages holds the potential to provide insights into the underlying inflammatory issues characteristic of AIDS and COVID-19 patients. Given the fundamental role of α7-nAChR in CAP functionality, it is conceivable that the observed alterations in expression are intricately linked to the inflammatory complications experienced by afflicted patients.

Grant/Funding Support. Funding for this research was provided by grant support: 5R25GM061151-20.

C.D. Walker1,2, O.R. Coulter1,2, T. Carter1,2, I. Parsons1, H.G. Sexton1,2, B.J. Henderson1, and M.L. Risher1,2
1Department of Biomedical Research, Joan C. Edwards School of Medicine, Marshall University, Huntington, WV, 2Hershel ‘Woody’ Williams Veterans Affairs Medical Center, Huntington, WV

Introduction/Background. Adolescence is a time of critical brain development and a time when alcohol consumption in the form of binge drinking is highly prevalent. Early onset binge drinking increases the risk of alcohol dependence and cognitive impairment later in life. Animal models have shown disruption of neuronal structure and function as a result of adolescent intermittent ethanol (AIE) exposure. However, the underlying mechanisms that elicit these changes are not well understood. Astrocytes have complex morphologies with extensive perisynaptic astrocyte processes (PAPs) that ensheathe pre- and post-synaptic terminals where they play an essential role in synaptic maintenance and signal transmission.

Hypothesis/Goal of Study. Here we investigate the consequences of AIE on astrocyte morphology, PAP-synaptic proximity, neuronal-to-astrocyte communication, and subsequent behavior. We hypothesize that AIE induces changes in astrocyte morphology resulting in a loss of PAP-synaptic proximity, reduced astrocyte Ca2+ responsivity to neuronal activation, and behavior. We further hypothesize that the behavioral deficits can be attenuated through the restoration of astrocyte Ca2+ signaling.

Methods and Results. Male Sprague Dawley rats received hippocampal intracranial injections of an astrocyte-specific adeno-associated virus containing green fluorescent protein (GFP), the astrocyte-specific Ca2+ indicator GCaMP6f, or hM3Dq (Gq) DREADDs. Beginning at postnatal day (PND) 30, animals received intermittent EtOH (5g/kg) or water via gavage over 16 days followed by a 26-day washout period. On PND72 acute hippocampal slices were prepared for immunohistochemistry (IHC, astrocyte morphology), electrophysiology (neuronal-astrocyte responsivity), or contextual fear conditioning. IHC was performed on GFP+ slices using the postsynaptic density marker PSD95. GFP+ astrocytes in the CA1 hippocampus were imaged using confocal microscopy and then reconstructed using IMARIS-Bitplane. Astrocyte Ca2+ activity was recorded from GCaMP6f+ CA1 hippocampal astrocytes following Schaffer Collateral stimulation. Contextual fear conditioning was completed using standard methods, pairing cue tone with foot shock following clozapine-n-oxide administration. Our data demonstrate that AIE results in a protracted decrease in astrocyte volume and PAP-synaptic colocalization that correlates with decreased astrocyte responsivity to neuronal activation as demonstrated by a reduction in astrocyte Ca2+ activity. When astrocyte Ca2+ activity was restored using Gq DREADDS, EtOH-induced behavioral deficits in the fear conditioning paradigm were attenuated.

Discussion/Conclusions. These findings demonstrate a critical role for astrocyte-neuronal communication in the regulation of behavior and contribute to our understanding of how adolescent EtOH disrupts neuronal-astrocyte communication, driving astrocytic and behavioral dysfunction that persists into adulthood.

Grant/Funding Support. Veterans Affairs Career Development Award BX002505 (MLR), Veterans Affairs Merit BX005403 (MLR), NIH-R21AA030086 (MLR), NASA West Virginia Space Grant Consortium, Training Grant NNX15AI01H (CDW)

Department of Biological Sciences, Northern Kentucky University, Highland Heights KY

Introduction/Background. The incidence of non-communicable inflammatory disorders is growing in the industrialized world. Recent research supports the theory that helminths and other commensalist (“helpful”) parasites can prevent and reduce symptoms of these disorders through alteration of the host immune system. Maternal helminths combined with helminth treatment at weaning rescues memory impairments in offspring that are caused by inflammation. We assessed maternal helminth status and its effects on cognitive function in offspring in a well-characterized model of neonatal infection.

Hypothesis/Goal of Study. Maternal helminths alone are sufficient to reduce neural inflammation following neonatal infection and the subsequent memory impairments for their adult offspring.

Methods and Results. Rat dams were treated with helminths 3 weeks prior to conception. On postnatal day 4, pups was treated with either PBS or live E. coli. As adults, offspring were trained on a hippocampal-dependent task, context pre-exposure fear conditioning. They were exposed to the novel context and injected with either saline or lipopolysaccharide (LPS). The following day they were shocked in the novel context, and on Day 3, they were tested for memory function by assessment of freezing behavior. Males and females differed in several behaviors, but maternal helminths did not rescue memory impairments. However, rats treated with helminths at weaning showed altered memory compared to controls. We also examined microglial immunohistochemistry and saw significant effects on the rate of microglial development in offspring born to helminth-treated mothers.

Discussion/Conclusions. Maternal helminths may have an effect during the early period of neural development in their offspring, but do not have lasting effects on offspring cognitive function. Helminths introduced at the time of weaning, however, have enduring effects on behavior into adulthood. Thus, the introduction of commensalist parasites as an intervention in inflammatory disorders is likely to require individual treatment rather than population-level preventative interventions.

Grant/Funding Support. NIH 1R15MH132060