Doctor of Philosophy, Miami University, 2024, Biology

Fragile X syndrome (FXS) is a common form of inherited intellectual disability and the primary monogenic cause of autism spectrum disorder. FXS results from a mutation to the Fragile X Mental Retardation 1 (FMR1) gene in the form of extra CGG repeats; greater than 200 repeats silences the gene and leads to the loss of the gene product, Fragile X Mental Retardation Protein (FMRP). As a neurodegenerative disorder, FXS research has primarily focused on the role of FMRP during development. Moreover, the role of FMRP in the innate immune system has not been well studied and differences between males and females are underexplored. The goal of this study was to better understand sex differences in fmr1 expression and function in the innate immune system using the cricket Acheta domesticus. To accomplish this, we first examined Ad-fmr1 expression in male and female neural and immune tissues before and after an immune challenge at sexually immature and sexually mature time points using quantitative real-time PCR. We found significant sex-, time-, and tissue-specific differences in Ad-fmr1 expression. Secondly, to examine FMRP's role in the innate immune system before and after a Serratia marcescens immune challenge, we used RNA interference to knockdown (KD) Ad-fmr1 expression of male and female crickets and examined the following innate immune parameters: survivorship, nodulation, fat body lysozyme expression, hemolymph lytic activity, nitric oxide synthase (nos) expression, and nitrite concentration. We found both sexes had increased lysozyme expression and lytic activity; nos expression was decreased while nitrite concentration decreased only in female hemolymph following frm1 KD. We also identified sex-specific differences for Ad-fmr1 KD crickets following an immune challenge. KD males were less likely to survive an immune challenge and did not nodulate bacteria as well as females. KD crickets had a significant increase in lysozyme expression, though only males had incre (open full item for complete abstract)

Committee: Kathleen Killian (Committee Chair); Paul James (Committee Member); Donghyung Lee (Committee Member); Haifei Shi (Committee Member); Dawn Blitz (Committee Member) Subjects: Biology

Doctor of Philosophy, Miami University, 2024, Biology

This dissertation investigates the sexually dimorphic expression of the nos gene, which encodes for the enzyme Nitric oxide synthase (NOS). NOS activation produces the gaseous signaling molecule nitric oxide (NO), which can impact immune function and behavior. Despite the crucial role of NO in insect physiology, the mechanisms underlying nos expression in both sexes and in different social environments remain poorly understood. Our primary aim was to elucidate the impact that a lack of social interaction can have on nos expression and immune function, providing new insights into insect physiology and behavior. We cloned and characterized the Ad-nos gene, revealing a close phylogenetic relationship with other orthopteran insects. Quantitative PCR analysis showed that early adult females exhibited significantly higher Ad-nos expression in nervous and thoracic fat body tissues than males, while males had higher expression in hemolymph. Following lipopolysaccharide (LPS) injection, males showed increased Ad-nos expression in brain and hemolymph, whereas females showed elevated expression in fat bodies and hemolymph. These findings suggest sex-specific immune strategies, with males prioritizing cellular immunity and females employing both humoral and cellular responses. We also examined the effects of social interaction during different developmental stages on NO levels and immune function. Comparing adult-isolated and nymph-isolated crickets, we found significant sex-dependent differences in Ad-nos expression. Nymph-isolated females had higher Ad-nos expression in brain and hemolymph, while nymph-isolated males showed higher expression in brain fat body and thoracic fat body tissues. NO activity assays indicated lower activity in nymph-isolated crickets compared to adult-isolated crickets. In addition, nymph-isolated females had fewer circulating hemocyte than males, while males had higher hemolymph protein content. Survival analysis post-LPS injection showed that adult (open full item for complete abstract)

Committee: Kathleen Killian (Advisor); Dawn Blitz (Committee Member); Jennifer Quinn (Committee Chair); Yoshinori Tomoyasu (Committee Member); Paul James (Committee Member) Subjects: Biology; Immunology; Molecular Biology; Neurosciences

Doctor of Philosophy, Miami University, 2019, Biology

Fragile X syndrome (FXS) is the most common form of inherited mental retardation and is the primary monogenetic cause of autism spectrum disorders. FXS is caused by a mutation in the regulatory region of the Fragile X Mental Retardation 1 (FMR1) gene, that ultimately leads to loss of the gene product Fragile X Mental Retardation Protein (FMRP). As FXS is a neurodevelopmental disorder, most studies have focused on the role of FMRP during development. However, a few studies have shown that the loss of FMRP during adulthood can impact learning, cognition, and behavior. The primary goal of this study was to decrease FMRP during adulthood and examine the impacts on immune function and social behavior using the cricket Acheta domesticus as a novel insect model. We examined immune function in males and females by assessing several important immune parameters including: the total number of circulating hemocytes in the hemolymph, the total hemolymph protein content, total phenoloxidase enzyme activity, and fat body lysozyme expression. We found that males and females exhibited similar changes in these immune parameters as a result of decreased Fmr1, but males were less likely to survive an immune challenge with an injection of lipopolysaccharide (LPS) from Serratia marcescens. This is the first study to demonstrate that decreased FMRP during adulthood causes sex-specific effects on the immune system. We also examined the effects of decreased Fmr1 on the agonistic behavior of adult male crickets. We observed that the average time to first physical contact during an agonistic interaction was significantly increased in fights between a control male and Fmr1 knockdown male and that Fmr1 knockdown males were less likely to initiate an aggressive interaction than control males. Despite initiating interactions less often, these males fought just as aggressively and won as often as controls. In this study, we also evaluated the parameters required to generate a successful parental R (open full item for complete abstract)

Committee: Kathleen Killian Dr. (Advisor) Subjects: Biology; Entomology; Genetics

Master of Science, Miami University, 2012, Zoology

The immune system is critical to an animal's survival. However, changes in immune function during insect development, and the factors that regulate these changes, are not well understood. Hemocytes carry out cellular defenses such as phagocytosis, nodulation, and encapsulation of pathogens. The enzyme phenoloxidase (PO) plays a key role in this process while the enzyme lysozyme breaks down bacterial cell walls. Total PO (TPA) and lysozyme activities, total blood hemocytes, and encapsulation ability were examined from late nymphal stages through early adulthood in the cricket, Acheta domesticus. TPA increased with age while encapsulation ability tended to decrease. The effects of crowding and light stress during nymphal development on adult immune parameters were determined. Crowding increased TPA in crickets housed in small but not large groups. Light stress negatively impacted survival, but not immune function. Further investigation of the effects of early life stress on adult immune function is thus warranted.

Committee: Kathleen A. Killian PhD (Advisor); Nancy G. Solomon PhD (Committee Member); Ann L. Rypstra PhD (Committee Member) Subjects:

Doctor of Philosophy, Miami University, 2005, Zoology

We examined the role of the cerci in relation to mating and escape in the cricket. Male cercal ablation significantly decreased mating success by reducing the ability of the male to hook the epiphallus onto the female subgenital plate and to transfer the spermatophore. We found that mechanosensory receptors located on the cerci of the male cricket supply important information on female position during spermatophore threading and transfer. Due to the role of cerci in both mating and escape, we next examined the behavioral switch that occurs that leads to mating. We mechanically stimulated crickets before, during, and after copulation. We found that most touch-evoked escape responses are suppressed in copulating males and females. The behavioral switch from escape to mating occurs following a males chemosensory contact with a female and requires the continued presence of the female. We identified the antennae as the primary source for chemosensory information with the maxillary palps being a secondary source. The loss of the female antennae had a significant negative effect on both female and male receptivity and mating behavior. Since the female antennae were found to stimulate the mating sequence in males, we next examined the role of external influences on the rhythmic bursting pattern that led to spermatophore threading. This rhythmic bursting pattern of action potential activity generated by genital motor neurons during spermatophore transfer was isolated to the terminal abdominal ganglion. Removal of both descending thoracic and abdominal inputs via complete transection of the ventral nerve cord and cercal sensory inputs by ablation of nerve 10d did not significantly affect this rhythmic activity. The ipsilateral cercal motor nerve and genital nerve remained coupled in their activities regardless of the input removed. Additionally, both the left and right genital nerves and the two cercal motor nerves also remained coupled. These results indicate that the rhythm (open full item for complete abstract)

Committee: Kathleen Killian (Advisor); Phyllis Callahan (Other); James Janik (Other); Nancy Solomon (Other) Subjects: Biology, Neuroscience