Milton, Sarah

The communication in the nervous system is a pharmacological balance between excitatory and inhibitory signals, and seizure behavior is one of the most common manifestations of when an imbalance occurs. Environmental toxins can cause significant disruption of excitation-inhibition balance, but while some toxins, like nerve agents, have known targets and require novel antidotes, some have unknown neurobiological mechanisms and require exploration. Of particular concern, there is little knowledge on how herbicides may affect neurological signaling. Glyphosate, the world’s most popular herbicide, was found to be in 80% of people’s urine, and since it is so prevalent, it is critical to understand its impact on both excitatory and inhibitory signaling. We used an electroshock assay developed for C. elegans to uncover evidence that glyphosate, and the commercial formula Roundup, disrupted the excitation-inhibition balance by blocking GABA-A receptors. This presented a novel hypothesis of an inhibitory neurobiological target for glyphosate. As glutamate is the major excitatory neurotransmitter in the human central nervous system, an electrophysiology assay using Drosophila was used and found that Roundup, but not glyphosate, reduced synaptic viability. This result directs attention to the undisclosed adjuvant component which may have a significant effect on synaptic transmission, though the exact mechanism requires further investigation.

As climate change threatens with sea-level rise and more storms, increased erosion could increase the need for beach nourishment. Alterations to sand characteristics may result in changes to the sea turtle nest microenvironment, impacting the temperature and oxygen levels which may affect hatchling performance. In this study, leatherback, loggerhead, and green nests were sampled from two sites with different sand characteristics in Juno Beach, Florida, USA. Gas exchange was higher in green turtle nests with a greater mixture of sediment. Darker sediment elevated nest temperatures. Finer sediment and a greater mixture of sediment in leatherback nests elevated the nest temperatures; conversely finer sediment, and a greater mixture of sediment decreased loggerhead and green nest temperatures. Elevated nest temperatures reduced leatherback, loggerhead, and green turtle hatchling performance. Understanding the relationships between beach composition, nest environment, and hatchling performance will aid management decisions essential to sea turtle conservation.

Under the expected warmer temperatures due to climate change, sea turtle embryos may be subjected to thermal conditions detrimental to nest success and hatchling quality; one trait which may be negatively affected is cognitive ability. In this study, loggerhead sea turtle eggs were acquired from Boca Raton, FL and lab incubated under two female-producing temperatures: an “optimal” temperature of 31°C and a sublethal temperature of 33°C. Cognitive ability of post-hatchlings, assessed via associative learning and reversal was investigated using a y-maze. The sublethal temperature decreased incubation duration, hatch success, hatchling growth rates and produced smaller hatchlings with significantly more scute anomalies. Hot hatchlings performed worse on the reversal, taking longer to train, and thus hint at an effect of incubation temperature on cognitive flexibility in loggerhead turtles. With temperatures rising on beaches in South Florida, this study provides evidence of further potential threats to hatchling quality and potentially even survival.

Climate change has the potential to expose sea turtle nests to higher temperatures, which may negatively impact sea turtle hatchling vigor. In this study, loggerhead and green hatchlings were sampled from the Boca Raton, Florida beach and via lab incubation, and hatchling vigor was determined. Elevated nest temperatures decreased loggerhead and green turtle hatchling performance and corticosterone levels, with the most significant effects found in hatchlings exposed to maximum incubation temperatures above 35°C during late development. Lab-incubated loggerhead post-hatchling corticosterone levels and growth rates were also determined. The differences seen in corticosterone levels with overall nest incubation temperatures, mean temperatures during early, middle or late stages of development, and its negative correlation with hatchling performance improves our understanding of the underlying physiological mechanisms linking elevated incubation temperatures and sub-lethal physiological effects that may significantly impact hatchling survival, a critical step for sea turtle conservation in south Florida and elsewhere.

The detrimental effects of oxidative stress caused by the accumulation of Reactive
Oxygen Species (ROS) have been acknowledged as major factors in aging, senescence and
several neurodegenerative diseases and conditions such as Parkinson’s disease and stroke
(ischemia/reperfusion). Mammalian models are extremely susceptible to these stresses that
follow the restoration of oxygen after anoxia; however, some organisms including the
freshwater turtle Trachemys scripta can withstand several bouts of anoxia and repeated
reoxygenation without any apparent pathology. T. scripta thus provides us with an
alternate vertebrate model in which we can investigate physiological mechanisms of
neuroprotection without the damaging effects that come with oxidative stress. The major
objective of this study was to investigate the protective mechanisms in the turtle brain
under conditions of anoxia and oxidative stress. Specifically, the focus is on the Methionine Sulfoxide Reductase system (Msr), an antioxidant and cellular repair system,
and how it is regulated to protect the brain against such stressors.
Previous studies in my lab have demonstrated that Msr mRNA and protein levels
are differentially upregulated during anoxia and reoxygenation. To investigate the
regulation of Msr, FOXO3a was directly induced by transfecting a human FOXO3a
plasmid into turtle brain cell cultures, as FOXO3a has been shown to regulate MsrA levels
in other animal models. Pharmacological manipulation of FOXO3a was also performed
using the green tea extract Epigallocatechin gallate (EGCG) as it has been shown to
increase expression of FOXO3a during oxidative stress conditions in other models. I found
that an induction of human FOXO3a increased FOXO3a levels and showed protection
against cell death during oxidative stress. Furthermore, treatment of cells with EGCG
increased expression of FOXO3a only when the cells were exposed to oxidative stress and
decreased cell death. Induction of FOXO3a and EGCG treatment did not increase MsrA
levels, however MsrB3 levels were upregulated under both treatments but only in the
presence of oxidative stress. These results suggest that MsrA and MsrB3 protect the cells
from oxidative stress damage through different molecular pathways and that EGCG may
be a therapeutic target to treat diseases related to damage by oxidative stress.