History of Previous Midlife Estradiol Treatment Permanently Alters Interactions of Brain Insulin-like Growth Factor-1 Signaling and Hippocampal Estrogen Synthesis to Enhance Cognitive Aging in a Rat Model of Menopause.

Across species, including humans, elevated levels of brain estrogen receptor (ER) α are associated with enhanced cognitive aging, even in the absence of circulating estrogens. In rodents, short-term estrogen treatment, such as that commonly used in the menopausal transition, results in long-term increases in ERα levels in the hippocampus, leading to enhanced memory long after termination of estrogen treatment. However, mechanisms by which increased levels of brain ERα enhances cognitive aging remain unclear. Here we demonstrate in aging female rats that insulin-like growth factor-1 (IGF-1), which can activate ER via ligand-independent mechanisms, requires concomitant synthesis of brain-derived neuroestrogens to phosphorylate ERα via MAPK signaling, ultimately resulting in enhanced memory. In a rat model of menopause involving long-term ovarian hormone deprivation, hippocampal neuroestrogen activity decreases, altering IGF-1 activity and resulting in impaired memory. However, this process is reversed by short-term estradiol treatment. Forty days of estradiol exposure following ovariectomy results in maintenance of neuroestrogen levels that persist beyond the period of hormone treatment, allowing for continued interactions between IGF-1 and neuroestrogen signaling, elevated levels of hippocampal ERα, and ultimately enhanced memory. Collectively, results demonstrate that short-term estradiol use following loss of ovarian function has long-lasting effects on hippocampal function and memory by dynamically regulating cellular mechanisms that promote activity of ERα in the absence of circulating estrogens. Translational impacts of these findings suggest lasting cognitive benefits of short-term estrogen use near menopause and highlight the importance of hippocampal ERα, independent from the role of circulating estrogens, in regulating memory in aging females.SIGNIFICANCE STATEMENT Declines in ovarian hormones following menopause coincide with increased risk of cognitive decline. Because of potential health risks, current recommendations are that menopausal estrogen therapy be limited to a few years. Long-term consequences for the brain and memory of this short-term midlife estrogen therapy are unclear. Here, in a rodent model of menopause, we determined mechanisms by which short-term midlife estrogen exposure can enhance hippocampal function and memory with cognitive benefits and molecular changes enduring long after termination of estrogen exposure. Our model indicates long-lasting benefits of maintaining hippocampal estrogen receptor function in the absence of ongoing estrogen exposure and suggests potential strategies for combating age-related cognitive decline.

Age Impacts the Burden That Reference Memory Imparts on an Increasing Working Memory Load and Modifies Relationships With Cholinergic Activity.

Rodent aging research often utilizes spatial mazes, such as the water radial-arm-maze (WRAM), to evaluate cognition. The WRAM can simultaneously measure spatial working and reference memory, wherein these two memory types are often represented as orthogonal. There is evidence, however, that these two memory forms yield interference at a high working memory load. The current study systematically evaluated whether the presence of a reference memory component impacts handling of an increasing working memory load. Young and aged female rats were tested to assess whether aging impacts this relationship. Cholinergic projections from the basal forebrain to the hippocampus and cortex can affect cognitive outcomes, and are negatively impacted by aging. To evaluate whether age-related changes in working and reference memory profiles are associated with cholinergic functioning, we assessed choline acetyltransferase activity in these behaviorally-tested rats. Results showed that young rats outperformed aged rats on a task testing solely working memory. The addition of a reference memory component deteriorated the ability to handle an increasing working memory load, such that young rats performed similar to their aged counterparts. Aged rats also had challenges when reference memory was present, but in a different context. Specifically, aged rats had difficulty remembering which reference memory arms they had entered within a session, compared to young rats. Further, aged rats that excelled in reference memory also excelled in working memory when working memory demand was high, a relationship not seen in young rats. Relationships between cholinergic activity and maze performance differed by age in direction and brain region, reflecting the complex role that the cholinergic system plays in memory and attentional processes across the female lifespan. Overall, the addition of a reference memory requirement detrimentally impacted the ability to handle working memory information across young and aged timepoints, especially when the working memory challenge was high; these age-related deficits manifested differently with the addition of a reference memory component. This interplay between working and reference memory provides insight into the multiple domains necessary to solve complex cognitive tasks, potentially improving the understanding of complexities of age- and disease- related memory failures and optimizing their respective treatments.

Inhibition of Estrogen Sulfotransferase (SULT1E1/EST) Ameliorates Ischemic Acute Kidney Injury in Mice.

BACKGROUND: Studies have suggested that estrogens may protect mice from AKI. Estrogen sulfotransferase (SULT1E1, or EST) plays an important role in estrogen homeostasis by sulfonating and deactivating estrogens, but studies on the role of SULT1E1 in AKI are lacking. METHODS: We used the renal ischemia-reperfusion model to investigate the role of SULT1E1 in AKI. We subjected wild-type mice, Sult1e1 knockout mice, and Sult1e1 knockout mice with liver-specific reconstitution of SULT1E1 expression to bilateral renal ischemia-reperfusion or sham surgery, either in the absence or presence of gonadectomy. We assessed relevant biochemical, histologic, and gene expression markers of kidney injury. We also used wild-type mice treated with the SULT1E1 inhibitor triclosan to determine the effect of pharmacologic inhibition of SULT1E1 on AKI. RESULTS: AKI induced the expression of Sult1e1 in a tissue-specific and sex-specific manner. It induced expression of Sult1e1 in the liver in both male and female mice, but Sult1e1 induction in the kidney occurred only in male mice. Genetic knockout or pharmacologic inhibition of Sult1e1 protected mice of both sexes from AKI, independent of the presence of sex hormones. Instead, a gene profiling analysis indicated that the renoprotective effect was associated with increased vitamin D receptor signaling. Liver-specific transgenic reconstitution of SULT1E1 in Sult1e1 knockout mice abolished the protection in male mice but not in female mice, indicating that Sult1e1's effect on AKI was also tissue-specific and sex-specific. CONCLUSIONS: SULT1E1 appears to have a novel function in the pathogenesis of AKI. Our findings suggest that inhibitors of SULT1E1 might have therapeutic utility in the clinical management of AKI.

Hepatic Estrogen Sulfotransferase Distantly Sensitizes Mice to Hemorrhagic Shock-Induced Acute Lung Injury.

Hemorrhagic shock (HS) is a potential life-threatening condition that may lead to injury to multiple organs, including the lung. The estrogen sulfotransferase (EST, or SULT1E1) is a conjugating enzyme that sulfonates and deactivates estrogens. In this report, we showed that the expression of Est was markedly induced in the liver but not in the lung of female mice subject to HS and resuscitation. Genetic ablation or pharmacological inhibition of Est effectively protected female mice from HS-induced acute lung injury (ALI), including interstitial edema, neutrophil mobilization and infiltration, and inflammation. The pulmonoprotective effect of Est ablation or inhibition was sex-specific, because the HS-induced ALI was not affected in male Est-/- mice. Mechanistically, the pulmonoprotective phenotype in female Est-/- mice was accompanied by increased lung and circulating levels of estrogens, attenuated pulmonary inflammation, and inhibition of neutrophil mobilization from the bone marrow and neutrophil infiltration to the lung, whereas the pulmonoprotective effect was abolished upon ovariectomy, suggesting that the protection was estrogen dependent. The pulmonoprotective effect of Est ablation was also tissue specific, as loss of Est had little effect on HS-induced liver injury. Moreover, transgenic reconstitution of human EST in the liver of global Est-/- mice abolished the pulmonoprotective effect, suggesting that it is the EST in the liver that sensitizes mice to HS-induced ALI. Taken together, our results revealed a sex- and tissue-specific role of EST in HS-induced ALI. Pharmacological inhibition of EST may represent an effective approach to manage HS-induced ALI.

Estradiol and selective estrogen receptor agonists differentially affect brain monoamines and amino acids levels in transitional and surgical menopausal rat models.

Estrogens have many beneficial effects in the brain. Previously, we evaluated the effects of two models of menopause (surgical vs. transitional) on multiple monoaminergic endpoints in different regions of the adult rat brain in comparison with levels in gonadally intact rats. Here we evaluated the effects of estrogen receptor (ER) agonist treatments in these same two models of menopause. Neurochemical endpoints were evaluated in the hippocampus (HPC), frontal cortex (FCX), and striatum (STR) of adult ovariectomized (OVX) rats and in rats that underwent selective and gradual ovarian follicle depletion by daily injection of 4-vinylcyclohexene-diepoxide (VCD), after 1- and 6-weeks treatment with 17ß-estradiol (E2), or with selective ERα (PPT), ERß (DPN), or GPR30 (G-1) agonists. Endpoints included serotonin (5-HT) and 5-Hydroxyindoleacetic acid, dopamine (DA), 3,4-Dihydroxyphenylacetic acid and homovanillic acid, norepinephrine (NE) and epinephrine, as well as the amino acids tryptophan (TRP) and tyrosine (TYR). Significant differences between the models were detected. OVX rats were much more sensitive to ER agonist treatments than VCD-treated rats. Significant differences between brain regions also were detected. Within OVX rats, more agonist effects were detected in the HPC than in any other region. One interesting finding was the substantial decrease in TRP and TYR detected in the HPC and FCX in response to agonist treatments, particularly in OVX rats. This is on top of the substantial decreases in TRP and TYR previously reported one week after OVX or VCD-treatments in comparison with gonadally intact controls. Other interesting findings included increases in the levels of 5-HT, DA, and NE in the HPC of OVX rats treated with DPN, increases in DA detected in the FCX of OVX rats treated with any of the ER agonists, and increases in 5-HT and DA detected in the STR of OVX rats treated with E2. Many effects that were observed after 1-week of treatment were no longer observed after 6-weeks of treatment, demonstrating that effects were temporary despite continued agonist treatment. Collectively, the results demonstrate significant differences in the effects of ER agonists on monoaminergic endpoints in OVX vs. VCD-treated rats that also were brain region-specific and time dependent. The fact that agonist treatments had lesser effects in VCD treated rats than in OVX rats may help to explain reports of lesser effects of estrogen replacement on cognitive performance in women that have undergone transitional vs. surgical menopause.

Detection of estradiol in rat brain tissues: Contribution of local versus systemic production.

Estrogens play important roles in regulating brain development, brain function, and behavior. Many studies have evaluated these effects using ovariectomized (OVX) rats or mice with different doses of estrogen replacement, assuming that estradiol levels in all regions of the brain are the same as levels achieved in the serum. It is well known, however, that the brain contains all the enzymes necessary to produce estrogens, and that estrogen levels in the brain are determined by both systemic and local production and are region-specific. The present study conducted a detailed analysis of the relationship between systemic levels of 17-ß-estradiol (E2) achieved by estrogen replacement and levels achieved in specific regions of the brain. Levels of E2 were measured in both brain and serum in OVX rats treated with different doses of estradiol benzoate (EB) using a novel and recently validated UPLC-MS/MS method. Results confirmed significantly higher levels of E2 in the brain than in serum in brain regions known to contain aromatase (ARO) activity, both in OVX controls and in rats treated with physiological doses of EB. Additional studies compared the level of E2 and testosterone (T) in the brain and serum between testosterone propionate (TP) treated OVX and male. This demonstrated higher levels of E2 in certain brain regions of males than in TP treated OVX females even though T levels in the brain and serum were similar between the two groups. Studies also demonstrated that the differences between serum and brain levels of E2 can be eliminated by letrozole (ARO inhibitor) treatment, which indicates that the differences are due to local ARO activity. Collectively the results provide a detailed analysis of brain region-specific E2 concentrations in OVX, E2-, and T-treated rats and demonstrate the degree to which these concentrations are ARO-dependent.

Effects of Cholinergic Lesions and Cholinesterase Inhibitors on Aromatase and Estrogen Receptor Expression in Different Regions of the Rat Brain.

Cholinergic projections have been shown to interact with estrogens in ways that influence synaptic plasticity and cognitive performance. The mechanisms are not well understood. The goal of this study was to investigate whether cholinergic projections influence brain estrogen production by affecting aromatase (ARO), or influence estrogen signaling by affecting estrogen receptor expression. In the first experiment, ovariectomized rats received intraseptal injection of the selective immunotoxin 192IgG-saporin to destroy cholinergic inputs to the hippocampus. In the second experiment ovariectomized rats received daily intraperitoneal injections of the cholinesterase inhibitors donepezil or galantamine for 1 week. ARO activity and relative levels of ARO, ERα, ERß, and GPR30 mRNAs were quantified in the hippocampus, frontal cortex, amygdala and preoptic area. Results show that the cholinergic lesions effectively removed cholinergic inputs to the hippocampus, but had no significant effect on ARO or on relative levels of ER mRNAs. Likewise, injections of the cholinesterase inhibitors had no effect on ARO or ER expression in most regions of the brain. This suggests that effects of cholinergic inputs on synaptic plasticity and neuronal function are not mediated by effects on local estrogen production or ER expression. One exception was the amygdala where treating with galantamine was associated with a significant increase in ARO activity. The amygdala is a key structure involved in registering fear and anxiety. Hence this finding may be clinically relevant to elderly patients who are treated for memory impairment and who also struggle with fear and anxiety disorders.

Comparison of transitional vs surgical menopause on monoamine and amino acid levels in the rat brain.

Loss of ovarian function has important effects on neurotransmitter production and release with corresponding effects on cognitive performance. To date, there has been little direct comparison of the effects of surgical and transitional menopause on neurotransmitter pathways in the brain. In this study, effects on monoamines, monoamine metabolites, and the amino acids tryptophan (TRP) and tyrosine (TYR) were evaluated in adult ovariectomized (OVX) rats and in rats that underwent selective and gradual ovarian follicle depletion by daily injection of 4-vinylcyclohexene-diepoxide (VCD). Tissues from the hippocampus (HPC), frontal cortex (FCX), and striatum (STR) were dissected and analyzed at 1- and 6-weeks following OVX or VCD treatments. Tissues from gonadally intact rats were collected at proestrus and diestrus to represent neurochemical levels during natural states of high and low estrogens. In gonadally intact rats, higher levels of serotonin (5-HT) were detected at proestrus than at diestrus in the FCX. In addition, the ratio of 5-hydroxyindoleacetic acid (5-HIAA)/5HT in the FCX and HPC was lower at proestrus than at diestrus, suggesting an effect on 5-HT turnover in these regions. No other significant differences between proestrus and diestrus were observed. In OVX- and VCD-treated rats, changes were observed which were both brain region- and time point-dependent. In the HPC levels of norepinephrine, 5-HIAA, TRP and TYR were significantly reduced at 1 week, but not 6 weeks, in both OVX and VCD-treated rats relative to proestrus and diestrus. In the FCX, dopamine levels were elevated at 6 weeks after OVX relative to diestrus. A similar trend was observed at 1 week (but not 6 weeks) following VCD treatment. In the STR, norepinephrine levels were elevated at 1 week following OVX, and HVA levels were elevated at 1 week, but not 6 weeks, following VCD treatment, relative to proestrus and diestrus. Collectively, these data provide the first comprehensive analysis comparing the effects of two models of menopause on multiple neuroendocrine endpoints in the brain. These effects likely contribute to effects of surgical and transitional menopause on brain function and cognitive performance that have been reported.

A microsomal based method to detect aromatase activity in different brain regions of the rat using ultra performance liquid chromatography-mass spectrometry.

Aromatase (ARO) is a cytochrome P450 enzyme that accounts for local estrogen production in the brain. The goal of this study was to develop a microsomal based assay to sensitively and reliably detect the low levels of ARO activity in different brain regions. Enzyme activity was detected based on the conversion of testosterone to estradiol. Quantity of estradiol was measured using ultra performance liquid chromatography-mass spectrometry. Detection was linear over a range of 2.5-200pg/ml estradiol, and was reproducible with intra- and inter-assay coefficients of variation (CV) <15%. Estradiol production using isolated microsomes was linear with time up to 30min as well as linearly related to amount of microsome. Substrate concentration curves revealed enzymatic kinetics (hippocampus: Vmax and Km: 0.57pmol estradiol/h per mg microsome and 48.58nM; amygdala: Vmax and Km: 1.69pmol estradiol/h per mg microsome and 48.4nM; preoptic area: Vmax and Km: 0.96pmol estradiol/h per mg microsome and 44.31nM) with testosterone used at a saturating concentration of 400nM. Anastrozole treatment blocked ARO activity in hippocampal and ovarian microsomes, indicating that the assay is specific for ARO. Also, we showed that the distribution of the long form ARO mRNA (CYP19A1) in different regions of the brain is correlated with ARO activity, with highest levels in the amygdala, followed by preoptic area and hippocampus. In the frontal cortex, very little long form ARO mRNA, and little to no ARO activity, were detected. These findings demonstrate that the microsomal incubation (MIB) assay is a sensitive and reliable method for quantifying ARO activity in discrete brain regions.

Targeted delivery of Doxorubicin by folic acid-decorated dual functional nanocarrier.

Doxorubicin (DOX) is one of the most commonly used antineoplastic agents, but its clinical application is oftentimes coupled with severe side effects. Selective delivery of DOX to tumors via nanosized drug carrier represents an attractive approach to this problem. Previously, we developed a dual functional nanomicellar carrier, PEG5K-embelin2 (PEG5K-EB2), which was able to deliver paclitaxel (PTX) selectively to tumors and to achieve an enhanced therapeutic effect. In the present study, we examined the utility of PEG5K-EB2 to deliver DOX to tumors. In addition, folic acid (FA) was coupled to the surface of the PEG5K-EB2 micelles (FA-PEG5K-EB2) to further improve the selective targetability of the system. DOX-loaded PEG5K-EB2 micelles were uniformly spherical particles with a diameter of approximately 20 nm. Incorporation of FA had minimal effect on the size of the particles. The DOX loading efficiency was as high as 91.7% and 93.5% for PEG5K-EB2 and FA-PEG5K-EB2, respectively. DOX formulated in PEG5K-EB2 micelles (with or without FA decoration) demonstrated sustained kinetics of DOX release compared to free DOX. FA-PEG5K-EB2 significantly facilitated the intracellular uptake of DOX over free DOX and PEGylated liposomal DOX (Doxil) in breast cancer cells, 4T1.2, and drug resistant cells, NCI/ADR-RES. P-gp ATPase assay showed that PEG5K-EB2 significantly inhibited the function of the P-gp efflux pump. The maximum tolerated dose of DOX-loaded PEG5K-EB2 micelles was 15 mg/kg in mice, which was 1.5-fold greater than that for free DOX. Pharmacokinetics (PK) and biodistribution studies showed that both types of DOX-loaded micelles, especially FA-PEG5K-EB2, were able to significantly prolong the blood circulation time of DOX and facilitate its preferential accumulation at the tumor tissue. Finally, DOX/PEG5K-EB2 mixed micelles demonstrated significantly enhanced tumor growth inhibitory effect with minimal toxicity in comparison to free DOX and Doxil and the antitumor activity was further enhanced after the decoration by folic acid. Our data suggest that FA-PEG5K-EB2 micelles represent a promising DOX delivery system that warrants more study in the future.

Estrogen sulfotransferase/SULT1E1 promotes human adipogenesis.

Estrogen sulfotransferase (EST/SULT1E1) is known to catalyze the sulfoconjugation and deactivation of estrogens. The goal of this study is to determine whether and how EST plays a role in human adipogenesis. By using human primary adipose-derived stem cells (ASCs) and whole-fat tissues from the abdominal subcutaneous fat of obese and nonobese subjects, we showed that the expression of EST was low in preadipocytes but increased upon differentiation. Overexpression and knockdown of EST in ASCs promoted and inhibited differentiation, respectively. The proadipogenic activity of EST in humans was opposite to the antiadipogenic effect of the same enzyme in rodents. Mechanistically, EST promoted adipogenesis by deactivating estrogens. The proadipogenic effect of EST can be recapitulated by using an estrogen receptor (ER) antagonist or ERα knockdown. In contrast, activation of ER in ASCs inhibited adipogenesis by decreasing the recruitment of the adipogenic peroxisome proliferator-activated receptor γ (PPARγ) onto its target gene promoters, whereas ER antagonism increased the recruitment of PPARγ to its target gene promoters. Linear regression analysis revealed a positive correlation between the expression of EST and body mass index (BMI), as well as a negative correlation between ERα expression and BMI. We conclude that EST is a proadipogenic factor which may serve as a druggable target to inhibit the turnover and accumulation of adipocytes in obese patients.

The decline in pulsatile GnRH release, as reflected by circulating LH concentrations, during the infant-juvenile transition in the agonadal male rhesus monkey (Macaca mulatta) is associated with a reduction in kisspeptin content of KNDy neurons of the arcuate nucleus in the hypothalamus.

Puberty in primates is timed by 2 hypothalamic events: during late infancy a decline in pulsatile GnRH release occurs, leading to a hypogonadotropic state that maintains quiescence of the prepubertal gonad; and in late juvenile development, pulsatile GnRH release is reactivated and puberty initiated, a phase of development that is dependent on kisspeptin signaling. In the present study, we determined whether the arrest of GnRH pulsatility in infancy was associated with a change in kisspeptin expression in the mediobasal hypothalamus (MBH). Kisspeptin was determined using immunohistochemistry in coronal hypothalamic sections from agonadal male rhesus monkeys during early infancy when GnRH release as reflected by circulating LH concentrations was robust and compared with that in juveniles in which GnRH pulsatility was arrested. The distribution of immunopositive kisspeptin neurons in the arcuate nucleus of the MBH of infants was similar to that previously reported for adults. Kisspeptin cell body number was greater in infants compared with juveniles, and at the middle to posterior level of the arcuate nucleus, this developmental difference was statistically significant. Neurokinin B in the MBH exhibited a similar distribution to that of kisspeptin and was colocalized with kisspeptin in approximately 60% of kisspeptin perikarya at both developmental stages. Intensity of GnRH fiber staining in the median eminence was robust at both stages. These findings indicate that the switch that shuts off pulsatile GnRH release during infancy and that guarantees the subsequent quiescence of the prepubertal gonad involves a reduction in a stimulatory kisspeptin tone to the GnRH neuronal network.

Incorporation of a selective sigma-2 receptor ligand enhances uptake of liposomes by multiple cancer cells.

BACKGROUND: The sigma-2 receptor is an attractive target for tumor imaging and targeted therapy because it is overexpressed in multiple types of solid tumors, including prostate cancer, breast cancer, and lung cancer. SV119 is a synthetic small molecule that binds to sigma-2 receptors with high affinity and specificity. This study investigates the utility of SV119 in mediating the selective targeting of liposomal vectors in various types of cancer cells. METHODS: SV119 was covalently linked with polyethylene glycol-dioleyl amido aspartic acid conjugate (PEG-DOA) to generate a novel functional lipid, SV119-PEG-DOA. This lipid was utilized for the preparation of targeted liposomes to enhance their uptake by cancer cells. Liposomes with various SV119 densities (0, 1, 3, and 5 mole%) were prepared and their cellular uptake was investigated in several tumor cell lines. In addition, doxorubicin (DOX) was loaded into the targeted and unmodified liposomes, and the cytotoxic effect on the DU-145 cells was evaluated by MTT assay. RESULTS: Liposomes with or without SV119-PEG-DOA both have a mean diameter of approximately 90 nm and a neutral charge. The incorporation of SV119-PEG-DOA significantly increased the cellular uptake of liposomes by the DU-145, PC-3, A549, 201T, and MCF-7 tumor cells, which was shown by fluorescence microscopy and the quantitative measurement of fluorescence intensity. In contrast, the incorporation of SV119 did not increase the uptake of liposomes by the normal BEAS-2B cells. In a time course study, the uptake of SV119 liposomes by DU-145 cells was also significantly higher at each time point compared to the unmodified liposomes. Furthermore, the DOX-loaded SV119 liposomes showed significantly higher cytotoxicity to DU-145 cells compared to the DOX-loaded unmodified liposomes. CONCLUSION: SV119 liposomes were developed for targeted drug delivery to cancer cells. The targeting efficiency and specificity of SV119 liposomes to cancer cells was demonstrated in vitro. The results of this study suggest that SV119-modified liposomes might be a promising drug carrier for tumor-targeted delivery.

Recognition of novel objects and their location in rats with selective cholinergic lesion of the medial septum.

The importance of cholinergic neurons projecting from the medial septum (MS) of the basal forebrain to the hippocampus in memory function has been controversial. The aim of this study was to determine whether loss of cholinergic neurons in the MS disrupts object and/or object location recognition in male Sprague-Dawley rats. Animals received intraseptal injections of either vehicle, or the selective cholinergic immunotoxin 192 IgG-saporin (SAP). 14 days later, rats were tested for novel object recognition (NOR). Twenty-four hours later, these same rats were tested for object location recognition (OLR) (recognition of a familiar object moved to a novel location). Intraseptal injections of SAP produced an 86% decrease in choline acetyltransferase (ChAT) activity in the hippocampus, and a 31% decrease in ChAT activity in the frontal cortex. SAP lesion had no significant effect on NOR, but produced a significant impairment in OLR in these same rats. The results support a role for septo-hippocampal cholinergic projections in memory for the location of objects, but not for novel object recognition.

Core competencies for research training in the clinical pharmaceutical sciences.

OBJECTIVE: To identify and apply core competencies for training students enrolled in the clinical pharmaceutical scientist PhD training program at the University of Pittsburgh School of Pharmacy. DESIGN: Faculty members reached consensus on the required core competencies for the program and mapped them to curricular and experiential requirements. ASSESSMENT: A rubric was created based on core competencies spanning 8 major categories of proficiency, and competencies of clinical versus traditional PhD training were delineated. A retrospective evaluation of the written comprehensive examinations of 12 former students was conducted using the rubric. Students scored above satisfactory in 11 out of 14 comprehensive examination metrics, with a mean score greater than 3.8 on a 5-point scale. CONCLUSIONS: The core competencies identified will provide an essential foundation for informed decision-making and assessment of PhD training in the clinical pharmaceutical sciences.

Evaluation of a combined therapeutic regimen of 8-OH-DPAT and environmental enrichment after experimental traumatic brain injury.

When provided individually, both the serotonin (5-HT(1A))-receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) and environmental enrichment (EE) enhance behavioral outcome and reduce histopathology after experimental traumatic brain injury (TBI). The aim of this study was to determine whether combining these therapies would yield greater benefit than either used alone. Anesthetized adult male rats received a cortical impact or sham injury and then were randomly assigned to enriched or standard (STD) housing, where either 8-OH-DPAT (0.1 mg/kg) or vehicle (1.0 mL/kg) was administered intraperitoneally once daily for 3 weeks. Motor and cognitive assessments were conducted on post-injury days 1-5 and 14-19, respectively. CA1/CA3 neurons and choline acetyltransferase-positive (ChAT(+)) medial septal cells were quantified at 3 weeks. 8-OH-DPAT and EE attenuated CA3 and ChAT(+) cell loss. Both therapies also enhanced motor recovery, acquisition of spatial learning, and memory retention, as verified by reduced times to traverse the beam and to locate an escape platform in the water maze, and a greater percentage of time spent searching in the target quadrant during a probe trial in the TBI + STD + 8-OH-DPAT, TBI + EE + 8-OH-DPAT, and TBI + EE + vehicle groups versus the TBI + STD + vehicle group (p ≤ 0.0016). No statistical distinctions were revealed between the TBI + EE + 8-OH-DPAT and TBI + EE + vehicle groups in functional outcome or CA1/CA3 cell survival, but there were significantly more ChAT(+) cells in the former (p = 0.003). These data suggest that a combined therapeutic regimen of 8-OH-DPAT and EE reduces TBI-induced ChAT(+) cell loss, but does not enhance hippocampal cell survival or neurobehavioral performance beyond that of either treatment alone. The findings underscore the complexity of combinational therapies and of elucidating potential targets for TBI.

Estrogen therapy and cognition: a review of the cholinergic hypothesis.

The pros and cons of estrogen therapy for use in postmenopausal women continue to be a major topic of debate in women's health. Much of this debate focuses on the potential benefits vs. harm of estrogen therapy on the brain and the risks for cognitive impairment associated with aging and Alzheimer's disease. Many animal and human studies suggest that estrogens can have significant beneficial effects on brain aging and cognition and reduce the risk of Alzheimer's-related dementia; however, others disagree. Important discoveries have been made, and hypotheses have emerged that may explain some of the inconsistencies. This review focuses on the cholinergic hypothesis, specifically on evidence that beneficial effects of estrogens on brain aging and cognition are related to interactions with cholinergic projections emanating from the basal forebrain. These cholinergic projections play an important role in learning and attentional processes, and their function is known to decline with advanced age and in association with Alzheimer's disease. Evidence suggests that many of the effects of estrogens on neuronal plasticity and function and cognitive performance are related to or rely upon interactions with these cholinergic projections; however, studies also suggest that the effectiveness of estrogen therapy decreases with age and time after loss of ovarian function. We propose a model in which deficits in basal forebrain cholinergic function contribute to age-related changes in the response to estrogen therapy. Based on this model, we propose that cholinergic-enhancing drugs, used in combination with an appropriate estrogen-containing drug regimen, may be a viable therapeutic strategy for use in older postmenopausal women with early evidence of mild cognitive decline.

Selective lesion of septal cholinergic neurons in rats impairs acquisition of a delayed matching to position T-maze task by delaying the shift from a response to a place strategy.

This study tested the hypothesis that septal cholinergic lesions impair acquisition of a delayed matching to position (DMP) T-maze task in male rats by affecting learning strategy. Rats received either the selective cholinergic immunotoxin, 192 IgG-saporin (SAP) or artificial cerebrospinal fluid directly into the medial septum. Two weeks later, animals were trained to acquire the DMP task. SAP-treated rats took significantly longer to acquire the task than corresponding controls. Both SAP-treated and control rats adopted a persistent turn and utilized a response strategy during early periods of training. By the time rats reached criterion the persistent turn was no longer evident, and all rats had shifted to an allocentric strategy, i.e., were relying on extramaze cues to a significant degree. During the acquisition period, SAP-treated rats spent significantly more days showing a persistent turn and using a response strategy than corresponding controls. The added time spent using a response strategy accounted entirely for the added days required to reach criterion among the SAP-treated rats. This suggests that the principal mechanism by which septal cholinergic lesions impair DMP acquisition in male rats is by increasing the predisposition to use a response vs. a place strategy, thereby affecting the ability to switch from one strategy to another.