Exposure to Homocysteic Acid Early in Postnatal Development Leads to a Mixed Depressive/Manic Behavioral Phenotype and Changes in NMDA Receptor Expression

Homocysteic acid (HCA), a NMDA receptor agonist, is an endogenous metabolite formed from the oxidation of homocysteine. Since hyperhomocysteinemia is a risk factor for several neuropsychiatric disorders, including bipolar disorder and major depressive disorder (MDD), we tested the hypothesis that elevated HCA levels in developing rats may induce alterations in NMDA receptor expression and the development of associated behaviors. Twenty postnatal male and twenty female rats were injected daily with either HCA or saline from P3 to P21. HCA-treated rats displayed increased risk-taking behavior, reduced social behavior, novelty-induced hyper-locomotion, anhedonia, and reduced spatial learning, consistent with a depressive state with manic tendencies. As expected, HCA treatment had no effect on motor coordination or paired-pulse inhibition. In addition to these behavioral changes, we observed that HCA led to an increase in expression of the GABAergic marker, GAD-67, in the cortex, but not the hippocampus, of both male and female rats. In the cortex, we also observed that HCA triggers a significant increase in the NMDAR2b:NMDAR2a subunit expression ratio in male rats, while female rats exhibited a decreased ratio. The results suggest that HCA triggered an increase in NMDAR2a expression in the hippocampus of both males and females. Finally, HCA also led to a decrease in NMDAR2b expression in females, but an increase in NMDAR2b expression in the hippocampus of males. We are currently examining the effect of HCA exposure on NMDAR1 subunit expression. Collectively, these data suggest that early postnatal exposure to HCA may lead to a mixed manic/depressive phenotype that may be accompanied by GABAergic signaling changes in the cortex. Given the proposed regulatory role of NMDA receptors on GABAergic interneuron activity and mood, we suggest that this may serve as a novel animal model for studies of complex mood disorders, such as bipolar disorder or MDD

Effect of Lithium Treatment on Homocysteic Acid-Induced Manic/Depressive Behaviors in Sprague-Dawley Rats

Bipolar disorder is a mood disorder for which the development of new treatments has been hindered by the lack of a reliable animal model. Previous studies in our lab have shown that daily injection of early postnatal rat pups with homocysteic acid (HCA), a homocysteine metabolite, leads to a mixed manic/depressive phenotype in adulthood. These results suggest that early HCA exposure may serve as a good model for bipolar disorder. To test the predictive validity of this model, we sought to determine if treatment of HCA-exposed animals with lithium was effective in reducing mixed manic/depressive behaviors elicited by HCA. As predicted, lithium treatment resulted in diminished manic-like behaviors in the HCA-treated rats and restored spatial learning in the Morris water maze of female, HCA-treated rats. Lithium also increased saccharine preference in female, HCA-treated animals and social activity in male, HCA-treated animals. However, lithium caused a reduction in saccharine preference and social behavior in the saline-treated rats. Finally, to determine if the effects of lithium were reversible, we removed the lithium from the rats’ diet for ten days and repeated the behavioral assessment. Removal of lithium resulted in the re-establishment of manic behavior in the elevated plus maze and a reduction of spatial learning in HCA-treated rats. However, the effects of lithium on the depression-associated behaviors of HCA-treated animals were not reversed, and all animals exhibited a strong increase saccharine preference and social behavior compared to when they were fed lithium. Thus, our results suggest that lithium is effective in treating the manic behavior, spatial learning deficits, anhedonia and reduced social interaction elicited by HCA as hypothesized. Collectively, these data provide further evidence that early postnatal exposure to HCA may serve as model for the mixed manic/depressive phenotype associated with bipolar disorder

Does Lithium Diminish Manic/Depressive Behaviors in Rats Associated with HCA Exposure?

The development of novel treatments for Bipolar Disorder has been hampered by the lack of a good animal model for the disease. The purpose of this study was to establish predictive validity for a novel animal model for bipolar disorder. Previous work in Dr. Chase’s lab has demonstrated that early exposure to the glutamatergic agonist, homocysteic acid (HCA), leads to a phenotype which is consistent with a mixed manic/depressive state as is often observed with bipolar disorder. Specifically, we observed that HCA-treated rats displayed increased risk-taking behavior, reduced social behavior, novelty-induced hyperlocomotion, anhedonia, and increased motivational behavior compared to control rats. Now we wish to determine if lithium, a common treatment for bipolar disorder, will lead to a reduction in depressive and manic behaviors associated with HCA. Therefore, we injected 23 rats daily with the endogenous, glutamatergic agonist homocysteic acid (HCA) from postnatal days 3-21. Another group of 23 rats received injection of saline. Six weeks later, the saccharine preference test, elevated plus maze, social interaction test, rotarod test, forced swim test and Morris Water Maze were conducted to assess anhedonia, risk taking behavior, social interaction, motor coordination, depressive behaviors, motivation and risk-taking behaviors. The rats were then treated with lithium for the remainder of the experiment and assessed in the same behavioral tests described previously to observe whether lithium was effective in reducing the behaviors associated with HCA exposure. If lithium is found to reduce manic and depressive behaviors in HCA-treated rats, this would suggest that the HCA model could potentially be used to identify novel pharmacological agents that would better treat the disorder

Isolation of 35 Mycobacteriophages and Genomic Analysis of the Novel Mycobacteriophage, Glass

Thirty-five new mycobacteriophages were isolated from soil samples collected on or nearby Hope College in Holland, Michigan. All were capable of infecting Mycobacterium smegmatis and produced a variety of plaque morphologies based on size, shape, and clarity, consistent with the isolation of an assortment of different phages. Both lytic and temperate phages appear represented in this collection. Purified phage stocks were used to prepare genomic DNA samples for restriction digest analysis. A comparison of those 35 digest results revealed few similarities among the group, further supporting our interpretation that most of the new phage isolates were distinct. One mycobacteriophage, Glass, was chosen for complete genome sequencing using the Illumina MiSeq platform and comparative genomic analysis. The predominant plaque produced by Glass at 32°C was turbid and 0.5-1.0mm in diameter, while plaque produced at 42°C was clear and 1.0-1.5mm in diameter. Genome sequence data for Glass revealed a relationship to a group of 12 mycobacteriophages in Cluster B2. The genome of Glass is 67.5 Kb, 69.0% GC, and contains 91 genes in agreement with the genome characteristics of closely related phage. A detailed analysis of the complete genome sequence and comparison with sequenced members of this small and unique group of mycobacteriophages is the subject of the second semester of this yearlong course and is presented

Isolation of 18 Novel Mycobacteriophages and Genomic Analyses of Krueger and Phrappuccino

Eighteen new mycobacteriophages were isolated from soil samples collected around the state of Michigan and parts of the United States. All phages were capable of infecting Mycobacterium smegmatis and were isolated through either enrichment or direct plating at 25°C. A variety of plaque morphologies were produced based on size, shape, and clarity; both lytic and temperate phages appear represented in this collection. Two mycobacteriophages, Krueger and Phrappuccino, were chosen for complete genome sequencing and comparative genomic analyses. The predominant plaque produced by Krueger at 32°C was circular and 2 mm in diameter. The predominant plaque produced by Phrappuccino at 32°C was 1 mm in diameter, and took 48 hours to appear. Complete genome sequence for Krueger revealed relationships to members of the novel Subcluster K6, while Phrappuccino was not closely related to any known phage and is currently classified as a Singleton. The genome of Krueger is 60.3 Kb, 66.5% GC, and contains 101 genes, including 1 tRNA(Lys-TTT) gene; the genome of Phrappuccino is 136.3 Kb, 67.4% GC, and contains 200 genes. While Phrappuccino is a Singleton, there is strong evidence at the morphological (Myoviridae) and genomic levels for a relationship to Cluster C phages. Despite this relationship, Phrappuccino does not carry any tRNA genes. Forty (39.6%) and thirty-six (18%) protein coding genes were assigned functions in Krueger and Phrappuccino, respectively, based on comparative analyses. A detailed analysis of the complete genome sequences and comparison with sequenced mycobacteriophages is the subject of the second semester of this yearlong course and is presented