Ionotropic Glutamate Receptors Show Unique Distribution and Localization in the Dorsal Cochlear Nucleus of the Rhesus Monkey

The dorsal cochlear nucleus (DCN) receives auditory information via the auditory nerve coming from the cochlea. It is responsible for much of the integration of auditory information, and it projects this auditory information to higher auditory brain centers for further processing. This study focuses on the DCN of adult Rhesus monkeys to characterize two specific cell types, the fusiform and cartwheel cell, based on morphometric parameters and type of glutamate receptor they express. The fusiform cell is the main projection neuron, while the cartwheel cell is the main inhibitory interneuron. Expression of AMPA glutamate receptor subunits is localized to certain cell types. The activity of the CN depends on the AMPA receptor subunit composition and expression. Immunocytochemistry, using specific antibodies for AMPA glutamate receptor subunits GluR1, GluR2/3 and GluR4, was used in conjunction with morphometry to determine the location, morphological characteristics and expression of AMPA receptor subunits in fusiform and cartwheel cells in the primate DCN. Qualitative as well as quantitative data indicates that there are important morphological differences in cell location and expression of AMPA glutamate receptor subunits between the rodent DCN and that of primates. GluR2/3 is widely expressed in the primate DCN. GluR1 is also widely expressed in the primate DCN. GluR4 is diffusely expressed. Expression of GluR2/3 and GluR4 in the primate is similar to that of the rodent. However, expression of GluR1 is different. GluR1 is only expressed by cartwheel cells in the rodent DCN, but is expressed by a variety of cells, including fusiform cells, in the DCN of the primate.

New aspects of estrogen action in the endometrium: Reciprocal interplay with retinoic acid pathway and a novel estrogen-regulated gene

The uterine endometrium is a major target for the estrogen. However, the molecular basis of estrogen action in the endometrium is largely unknown. I have used two approaches to study the effects of estrogen on the endometrium. One approach involved the study of the interaction between estrogen and retinoic acid (RA) pathways in the endometrium. I have demonstrated that estrogen administration to rodents and estrogen replacement therapy (ERT) in postmenopausal women selectively induced the endometrial expression of retinaldehyde dehydrogenase II (RALDH2), a critical enzyme of RA biosynthesis. RALDH2 was expressed exclusively in the stromal cells, especially in the stroma adjacent to the luminal and glandular epithelia. The induction of RALDH2 by estrogen required estrogen receptor and occurred via a direct increase in RALDH2 transcription. Among the three RA receptors, estrogen selectively induced the expression of RARα. In parallel, estrogen also increased the utilization of all-trans retinol (the substrate for RA biosynthesis) and the expression of two RA-regulated marker genes, cellular retinoic acid binding protein II (CRABP2) and tissue transglutaminase (tTG) in the endometrium. Thus estrogen coordinately upregulated both the production and signaling of RA in both the rodent and human endometrium. This coordinate upregulation of RA system appeared to play a role in counterbalancing the stimulatory effects of estrogen on the endometrium, since the depletion of endogenous RA in mice led to an increase in estrogen-stimulated stromal proliferation and endometrial Akt phosphorylation. In addition, I have also used a systematic approach (DNA microarray) to categorize genes and pathways affected by the ERT in the endometrium of postmenopausal women and identified a novel estrogen-regulated gene EIG121. EIG121 was exclusively expressed in the glandular epithelial cells of the endometrium and induced by estrogen in vivo and in cultured cell lines. Compared with the normal endometrium, EIG121 was highly overexpressed in type 1 endometrial cancer, but profoundly suppressed in type 2 endometrial tumors. Taken together, these studies suggested that estrogen regulates the expression of many genes of both the pro-proliferative and anti-proliferative pathways and the abnormality of these pathways may increase the risks for estrogen-dependent endometrial hyperplasia and endometrial cancer. ^

Behavioral Implications of Knockout for the Dyslexia-Risk Gene Dcdc2 in Mice

Several genetic linkage and epidemiological studies have provided strong evidence that DCDC2 is a candidate gene for developmental dyslexia, a disorder that impairs a person’s reading ability despite adequate intelligence, education, and socio-economic status. Studies investigating embryonic intra-ventricular RNA interference (RNAi) of Dcdc2, a rat homolog of the DCDC2 gene in humans, indicate disruptions in neuronal migration in the rat cortex during development. Interestingly, these anatomical anomalies are consistent with post mortem histological analysis of human dyslexic patients. Other rodent models of cortical developmental disruption have shown impairment in rapid auditory processing and learning maze tasks in affected subjects. The current study investigates the rapid auditory processing abilities of mice heterozygous for Dcdc2 (one functioning Dcdc2 allele) and mice with a homozygous knockout of Dcdc2 (no functioning Dcdc2 allele). It is important to note that this genetic model for behavioral assessment is still in the pilot stage. However, preliminary results suggest that mice with a genetic mutation of Dcdc2 have impaired rapid auditory processing, as well as non-spatial maze learning and memory ability, as compared to wildtypes. By genetically knocking out Dcdc2 in mice, behavioral features associated with Dcdc2 can be characterized, along with other neurological abnormalities that may arise due to the loss of the functioning gene.

Investigation of the molecular mechanisms of endometrial hyperplasia

While there is considerable information on the molecular aberrations associated with the development of endometrial cancer, very little is known of changes in gene expression associated with its antecedent premalignant condition, endometrial hyperplasia. In order to address this, we have compared the level of expression of components of the IGF-I signaling pathway in human endometrial hyperplasia to their level of expression in both the normal pre-menopausal endometrium and endometrial carcinoma. We have also characterized the molecular characteristics of endometrial hyperplasia as it occurs in a murine model of hormone-dependent tumorigenesis of the female reproductive tract. ^ There was a significant and selective increase in the expression of the IGF-I Receptor (IGF-IR) in both human hyperplasia and carcinoma as compared to the normal endometrium. The receptor was also activated, as judged by increased tyrosine phosphorylation. In addition, in hyperplasia and carcinoma there is activation of the downstream component Akt. The expression of the PTEN tumor suppressor is decreased in a subset of subjects with hyperplasia and in all of the carcinomas. The simultaneous loss of PTEN expression and increased IGF-IR activation in the hyperplastic endometrium was associated with an increased incidence of endometrial carcinoma elsewhere within the uterus. In the rodent hyperplasia, there was a significant increase in the expression and activation of Akt that appears to be attributable to a marked increase in the expression of IGF-II. ^ Our studies have demonstrated the pathologic proliferation of both the human and rodent endometrium is linked to a marked activation of the Akt pathway. However the cause of this dysregulation is different in the human disease and the animal model. In rodents, hyperplasia is linked to increased expression of one of the ligands of the IGF-IR, IGF-II. In humans the IGF-I receptor itself is upregulated and activated. Additional activation of the Akt pathway via the suppression of PTEN activity, results in conditions that are associated with the marked increase in the probability of developing endometrial cancer. Our data suggests that increased activity of the IGF-I pathway plays the key role in the hyperproliferative state characteristic of endometrial hyperplasia and cancer.^

New insights into the role of the duodenal vitamin D receptor in calcium homeostasis

It is generally believed that 1,25(OH)2D3, bound to its receptor (VDR) contributes to calcium homeostasis by regulating active calcium absorption in the proximal small intestine. However, studying patients with hereditary vitamin D-resistant rickets (HVDRR) provided investigators with a better understanding of VDR's role in calcium homeostasis. HVDRR patients have inactivating mutations in the VDR, and as a consequence they develop hypocalcemia, hyperparathyroidism and severe rickets. However, these phenotypes can be corrected if the patients are given IV infusions of calcium or dietary calcium. This raises the question of what is the physiological significance of VDR-regulated active calcium absorption if calcium homeostasis can be restored independently of the VDR. ^ In order to distinguish the contribution of VDR in the proximal small intestine to overall calcium homeostasis, I generated transgenic mice expressing the human VDR (hVDR) exclusively in the proximal small intestine of mVDR-/- mice by using an hVDR-expressing transgene driven by the duodenal-specific adenosine deaminase enhancer (hVDR+/mVDR-/-). hVDR+/mVDR-/- mice expressed transcriptionally active hVDR only in the proximal small intestine and responded to 1,25(OH)2D3 by up-regulating expression of TRPV6 and calbindin D9K, genes involved in calcium absorption. Furthermore, ligated duodenal loop assays determined that calcium absorption in hVDR+/mVDR-/- mice was as responsive to 1,25(OH)2D3 as in WT mice. Despite having a functional hVDR in the proximal small intestine, hVDR+/mVDR-/- mice were hypocalcemic, had hyperparathyroidism, and were rachitic when fed a normal rodent diet at weaning, as were the mVDR-/- mice. However, when fed a high calcium, phosphorus, and lactose diet (rescue diet), the hVDR+/mVDR-/- mice responded more effectively than the mVDR-/- mice by down-regulation of parathyroid hormone production and by a greater increase in bone mineralization. Furthermore, when three-month-old rachitic mice were fed a rescue diet for 3 weeks, serum calcium and bone mineral content were normalized in hVDR+/mVDR-/- mice, but not in mVDR-/- mice. ^ In conclusion, hVDR expression enabled young mice to better use the rescue diet than mVDR-/- mice. Expression of transgenic hVDR also protected the ability of older mice to respond to the rescue diet despite the absence of the VDR elsewhere in the intestinal tract. I propose that because hVDR+/mVDR-/- mice responded better than mVDR-/- mice to the rescue diet, it is likely that VDR expression in the proximal small intestine is necessary in nutritional (insufficient dietary calcium) and physiological (age) conditions when passive calcium absorption is inadequate. ^

A Pre-Clinical Assessment of Minocycline for Treatment of Chronic Neuropathic Pain after Spinal Cord Injury

Patients living with a spinal cord injury (SCI) often develop chronic neuropathic pain (CNP). Unfortunately, the clinically approved, current standard of treatment, gabapentin, only provides temporary pain relief. This treatment can cause numerous adverse side effects that negatively affect the daily lives of SCI patients. There is a great need for alternative, effective treatments for SCI-dependent CNP. Minocycline, an FDA-approved antibiotic, has been widely prescribed for the treatment of acne for several decades. However, recent studies demonstrate that minocycline has neuroprotective properties in several pre-clinical rodent models of CNS trauma and disease. Pre-clinical studies also show that short-term minocycline treatment can prevent the onset of CNP when delivered during the acute stage of SCI and can also transiently attenuate established CNP when delivered briefly during the chronic stage of SCI. However, the potential to abolish or attenuate CNP via long-term administration of minocycline after SCI is unknown. The purpose of this study was to investigate the potential efficacy and safety of long-term administration of minocycline to abolish or attenuate CNP following SCI. A severe spinal contusion injury was administered on adult, male, Sprague-Dawley rats. At day 29 post-injury, I initiated a three-week treatment regimen of daily administration with minocycline (50 mg/kg), gabapentin (50 mg/kg) or saline. The minocycline treatment group demonstrated a significant reduction in below-level mechanical allodynia and above- level hyperalgesia while on their treatment regimen. After a ten-day washout period of minocycline, the animals continued to demonstrate a significant reduction in below-level mechanical allodynia and above-level hyperalgesia. However, minocycline-treated animals exhibited abnormal weight gain and hepatotoxicity compared to gapabentin-treated or vehicle-treated subjects.The results support previous findings that minocycline can attenuate CNP after SCI and suggested that minocycline can also attenuate CNP via long-term delivery of minocycline after SCI (36). The data also suggested that minocycline had a lasting effect at reducing pain symptoms. However, the adverse side effects of long-term use of minocycline should not be ignored in the rodent model. Gabapentin treatment caused a significant decrease in below-level mechanical allodynia and below-level hyperalgesia during the treatment regimen. Because gabapentin treatment has an analgesic effect at the concentration I administered, the results were expected. However, I also found that gabapentin-treated animals demonstrated a sustained reduction in pain ten days after treatment withdrawal. This result was unexpected because gabapentin has a short half-life of 1.7 hours in rodents and previous studies have demonstrated that pre-drug pain levels return shortly after withdrawal of treatment. Additionally, the gabapentin-treated animals demonstrated a significant and sustained increase in rearing events compared with all other treatment groups which suggested that gabapentin treatment was not only capable of reducing pain long-term but may also significantly improve trunk stability or improve motor function recovery.

Mechanisms of adenovirus 5 E1A -mediated tumor suppression and E1A-mediated apoptosis

The adenovirus type 5 E1A gene products have numerous functions in cells, which serve as useful tools in studying the mechanisms of either oncogenesis or tumor suppression. To understand the mechanisms of E1A-mediated tumor suppression, we introduced an Ad5 E1A gene into murine melanoma cells, and characterized E1A-mediated biological functions both in vitro and in vivo. The results of the study indicated that: (i) Ad5 E1A mediated tumor suppression in rodent tumor cells; (ii) E1A-mediated tumor suppression is associated with E1A-mediated apoptosis in vivo.^ To determine which functional region(s) of E1A is(are) required for E1A-mediated apoptosis and whether E1A-mediated apoptosis is required for E1A-mediated tumor suppression, we established stable transfectants of E1A mutants, which have deletion mutation at either the N-terminal (p300-binding) or the CR2 (pRb-binding) domain or both, and then characterized biological functions both in vitro and in vivo. The results of the study indicate that the CR2 domain of E1A is required for E1A-mediated apoptosis, while the N-terminal domain of E1A is dispensable. Interestingly, either of the two domains is able to mediate tumor suppression, since mutant E1A with a single deletion at either domain still suppressed tumor growth. Importantly, deletion mutations at both the N-terminal and the CR2 domains of E1A abrogated E1A-mediated tumor suppression, suggesting both regions are required for E1A-mediated tumor suppression. The results demonstrate that E1A-mediated apoptosis is not the only mechanism for E1A-mediated tumor suppression. Thus, the N-terminal and CR2 domains of E1A mediated two independent mechanisms of tumor suppression.^ To understand the mechanism of E1A-mediated apoptosis, we examined the temporal relationship of molecular events during the apoptotic cascades after UV radiation and serum depletion in both the E1A-expressing cells and parental cells. Kinetic analysis of JNK activity indicates that the JNK pathway is greatly increased in response to UV light in E1A transfectants, suggesting that extracellular stress stimuli have been converted into intracellular stress signals with greater magnitude in E1A transfectants than those in parental cells. Thus, E1A-mediated sensitization precedes these events. As ceramide has been proposed as second messenger and upstream activator of JNK pathway for stress-induced apoptosis, we also examined the roles of ceramide in apoptosis and the relationship with JNK pathway. The results indicate that E1A transfectants do not have increased sensitivity to ceramide. Therefore, E1A-mediated sensitization to UV radiation cannot be attributed to an increased sensitivity to ceramide. Furthermore, UV-induced JNK activation correlates with UV-induced apoptosis, while lethal dose of ceramide does not activate JNK. Thus, activation of JNK pathway is independent of the ceramide pathway. In addition, E1A transfectants also have increased activation of NF-kB in response to UV. These results suggest that E1A-mediated sensitization is an early event which associates with conversion of extracellular stress stimuli into amplified intracellular signals. The mechanism of E1A-mediated sensitization and its relationship with other pathways are discussed. ^

Geochemistry of recent sediments from the Indian Ocean

During the International Indian Ocean Expedition (1964/65) sediment cores were taken on six profiles off the western coast of the Indian Subcontinent. These profiles run approximately perpendicular to the coast, from the deep-sea over the continental slope to the continental shelf. Additional samples and cores were taken in a dense pattern in front of the delta of the Indus River. This pattern of sampling covered not only marine sediments, but also river and beach sediments in Pakistan. The marine samples were obtained with piston, gravity and box corers and by a Van Veen grab sampler. The longest piston core is about 5 meters long. 1. Distribution of the elements on the sediment surface The area of maximal carbonate values (aprox. 80-100% CaCO3) essentially coincides with the continental shelf. The highest Sr values were observed largely within this area, but only in the vicinity of the Gulf of Cambay. Mainly the aragonitic coprolites are responsible for those high Sr contents. The Mg contents of the carbonates are comparatively low; surprisingly enough the highest Mg concentrations were also measured in the coprolites. The maximum contents of organic matter (Core) were found along the upper part of the continental slope. They coincide with the highest porosity and water content of the sediments. Frequently the decomposition of organic matter by oxydation is responsible for the measured Corg contents. On the other side the quantity of originally deposited organic material is less important in most cases. The enrichment of the "bauxitophile" elements Fe, Ti, Cr and V in the carbonate- and quartz-free portions of the sediments is essentially due to the influence of coarse terrigenous detritus. For the elements Mn, Ni and Cu (in per cent of the carbonateand quartz-free sediment) a strong enrichment was observed in the deep-sea realm. The strong increase in Mn toward the deep-sea is explained by authigenesis of Mn-Fe-concretions. Mn-nodules form only under oxydizing conditions which obviously are possible only at very low rates of deposition. The Mg, B and, probably also Mn contents in the clay minerals increase with increasing distance from the continent. This can be explained by the higher adsorption of those elements from sea water because of increasing duration of the clay mineral transport. The comparison of median contents of some elements in our deep-sea samples with deep-sea sediments described by TUREKIAN & WEDEPOHL (1961) shows that clear differences in concentration exist only in the case of "bauxitophile" elements Cr and Be. The Cr and Be contents show a clear increase in the Indian Ocean deep-sea samples compared to those described by TUREKIAn & WEDEPOHL (1961) which can obviously be attributed to the enrichment in the lateritic and bauxitic parent rocks. The different behaviour of the elements Fe, Ti and Mn during decomposition of the source rocks, transport to the sea and during oxydizing and reducing conditions in the marine environment can be illustrated by Ti02/Fe and MnO/Fe ratios. The different compositions of the sediments off the Indus Delta and those of the remaining part of the area investigated are characterized by a different distribution of the elements Mn and Ti. 2. Chemical inhomogenities in the sediments Most longer cores show 3 intervals defined by chemical and sedimentological differences. The top-most interval is coarse-grained, the intermedial interval is fine grained and the lower one again somewhat coarser. At the same time it is possible to observe differences from interval to interval in the organogenic and detrital constituents. During the formation of the middle interval different conditions of sedimentation from those active during the previous and subsequent periods have obviously prevailed. Looking more closely at the organogenic constituents it is remarkable that during the formation of the finer interval conditions of a more intensive oxydation have prevailed that was the case before and after: Core decreases, whereas P shows a relative increase. This may be explained by slower sedimentation rate or by a vertical migration of the oxygen rich zone of the sea-water. The modifications of the elements from minerals in detrital portion of the sediments support an explanation ascribing this fact to modifications of the conditions of denudation and transportation which can come about through a climatic change or through tectonic causes. The paleontological investigations have shown (ZOBEL, in press) that in some of the cores the middle stratum of fine sedimentation represents optimal conditions for organic life. This fact suggests also oxydizing conditions during the sedimentation of this interval. In addition to the depositional stratification an oxydation zone characterized by Mn-enrichment can be recognized. The thickness of the oxidation zone decreases towards the coast and thins out along the middle part of the continental slope. At those places, where the oxydation zone is extremely thin, enrichment of Mn has its maximum. This phenomenon can probably be attributed to the migration of Mn taking place in its dissociated form within the sediment under reducing conditions. On the other side this Mn-migration in the sediment does not take place in the deep-sea, where oxydizing conditions prevail. 3. Interstitial waters in the sediments Already at very small core depths, the interstitial waters have undergone a distinct modification compared with the overlying sea water. This distinct modification applies both to total salinity and to the individual ions. As to the beginning of diagenesis the following conclusions can be drawn: a) A strong K-increase occurs already at an early stage. It may be attributable to a diffusion barrier or to an exchange of Mg-ions on the clays. Part of this increase may also originate from the decomposition of K-containing silicates (mica and feldspars). A K-decrease owing to the formation of illite (WEAVER 1967), however, occurs only at much greater sediment depth. b) Because of an organic protective coating, the dissolution of carbonate is delayed in recent organogenic carbonates. At the same time some Ca is probably being adsorbed on clay minerals. Consequently the Ca-content of the interstitial water drops below the Ca-content of the sea water. c) Already at an early stage the Mg adsorption on the clays is completed. The adsorbed Mg is later available for diagenetic mineral formations and transformations.

Avian species identity and predation success in tropical cacao agroforestry of the Napu Valley in Central Sulawesi, Indonesia

Avian ecosystem services such as the suppression of pests are considered being of high ecological and economic importance in a range of ecosystems, especially in tropical agroforestry. But how bird predation success is related to the diversity and composition of the bird community, as well as local and landscape factors, is poorly understood. The author quantified arthropod predation in relation to the identity and diversity of insectivorous birds, using experimental exposure of artificial, caterpillar-like prey on smallholder cacao agroforestry systems, differing in local shade management and distance to primary forest. The bird community was assessed using both mist netting (targeting on active understory insectivores) and point count (higher completeness of species inventories) sampling. The study was conducted in a land use dominated area in Central Sulawesi, Indonesia, adjacent to the Lore Lindu National Park. We selected 15 smallholder cacao plantations as sites for bird and bat exclosure experiments in March 2010. Until July 2011, we recorded several data in this study area, including the bird community data, cacao tree data and bird predation experiments that are presented here. We found that avian predation success can be driven by single and abundant insectivorous species, rather than by overall bird species richness. Forest proximity was important for enhancing the density of this key species, but did also promote bird species richness. The availability of local shade trees had no effects on the local bird community or avian predation success. Our findings are both of economical as well as ecological interest because the conservation of nearby forest remnants will likely benefit human needs and biodiversity conservation alike.

Leftovers in seed dispersal: ecological implications of partial seed consumption for oak regeneration

1. Successful seed dispersal by animals is assumed to occur when undamaged seeds arrive at a favourable microsite. Most seed removal and dispersal studies consider only two possible seed fates, predation or escape intact. Whether partial consumption of seeds has ecological implications for natural regeneration is unclear. We studied partial consumption of seeds in a rodent-dispersed oak species. 2. Fifteen percent of dispersed acorns were found partially eaten in a field experiment. Most damage affected only the basal portion of the seeds, resulting in no embryo damage. Partially eaten acorns had no differences in dispersal distance compared to intact acorns but were recovered at farther distances than completely consumed acorns. 3. Partially eaten acorns were found under shrub cover unlike intact acorns that were mostly dispersed to open microhabitats. 4. Partially eaten acorns were not found buried proportionally more often than intact acorns, leading to desiccation and exposure to biotic agents (predators, bacteria and fungi). However, partial consumption caused more rapid germination, which enables the acorns to tolerate the negative effects of exposure. 5. Re-caching and shrub cover as microhabitat of destination promote partial seed consumption. Larger acorns escaped predation more often and had higher uneaten cotyledon mass. Satiation at seed level is the most plausible explanation for partial consumption. 6. Partial consumption caused no differences in root biomass when acorns experienced only small cotyledon loss. However, root biomass was lower when acorns experienced heavy loss of tissue but, surprisingly, they produced longer roots, which allow the seeds to gain access sooner to deeper resources. 7.Synthesis. Partial consumption of acorns is an important event in the oak regeneration process, both quantitatively and qualitatively. Most acorns were damaged non-lethally, without decreasing both dispersal distances and the probability of successful establishment. Faster germination and production of longer roots allow partially eaten seeds to tolerate better the exposure disadvantages caused by the removal of the pericarp and the non-buried deposition. Consequently, partially consumed seeds can contribute significantly to natural regeneration and must be considered in future seed dispersal studies.

Disentangling factors controlling fruit and seed removal by rodents in temperate forests

Fleshy fruits fall on to the ground together with cleaned seeds previously ingested by primary dispersers, offering a wide range of fruits and seeds to the ground foragers. Although nutritional properties strongly differ between fruits and seeds, this different seed presentation (cleaned seeds versus seeds within the pulp) has not been addressed in seed removal studies. This study reports on the removal of fruits versus their seeds in five fleshy-fruited species in a temperate forest. We found that rodents removed most of the seeds and partially consumed most of the fruits, preferring seeds to fruits. Rodents bit the fruits to extract the seeds, leaving most of the pulp. We found a preference ranking for the seeds (Sorbus aucuparia>Ilex aquifolium>Sorbus aria>Rosa canina>Crataegus monogyna) but no preferences were found for the fruits, probably due to their similarities in pulp constituents. Seed and fruit choice were affected by chemical and physical properties and not by their size. The presence of alternative and preferred seeds (nuts) delayed the encounter of the fruits and seeds and diminished their removal rates. We found that higher rodent abundance is not necessarily associated with higher removal rates of fleshy fruits. Rodent abundance, fruit size and seed size are minor factors in the removal of fleshy fruits and their seeds. This study underlines that scatter-hoarding rodents are important removers of fleshy fruits and their seeds, producing a differential seed removal depending on the seed presentation (with or without pulp), the nutritional properties of the seeds (but not of the fruits) and the presence of alternative food

Moonlight and shelter cause differential seed selection and removal by rodents

Various environmental factors may influence the foraging behaviour of seed dispersers which could ultimately affect the seed dispersal process. We examined whether moonlight levels and the presence or absence of rodentshelter affect rodentseedremoval (rate, handling time and time of removal) and seedselection (size and species) among seven oak species. The presence or absence of safe microhabitats was found to be more important than moonlight levels in the removal of seeds. Bright moonlight caused a different temporal distribution of seedremoval throughout the night but only affected the overall removal rates in open microhabitats. Seeds were removed more rapidly in open microhabitat (regardless of the moon phase), decreasing the time allocated to seed discrimination and translocation. Only in open microhabitats did increasing levels of moonlight decrease the time allocated to selection and removal of seeds. As a result, a more precise seedselection was made under shelter, owing to lower levels of predation risk. Rodent ranking preference for species was identical between full/new moon in shelter but not in open microhabitats. For all treatments, species selection by rodents was much stronger than size selection. Nevertheless, heavy seeds, which require more energy and time to be transported, were preferentially removed under shelter, where there is no time restriction to move the seeds. Our findings reveal that seedselection is safety dependent and, therefore, microhabitats in which seeds are located (sheltered versus exposed) and moonlight levels in open areas should be taken into account in rodent food selection studies.

Colocalization of α-actinin and Synaptopodin in the Pyramidal Cell Axon Initial Segment

The cisternal organelle that resides in the axon initial segment (AIS) of neocortical and hippocampal pyramidal cells is thought to be involved in regulating the Ca(2+) available to maintain AIS scaffolding proteins, thereby preserving normal AIS structure and function. Through immunocytochemistry and correlative light and electron microscopy, we show here that the actin-binding protein ?-actinin is present in the typical cistenal organelle of rodent pyramidal neurons as well as in a large structure in the AIS of a subpopulation of layer V pyramidal cells that we have called the "giant saccular organelle." Indeed, this localization of ?-actinin in the AIS is dependent on the integrity of the actin cytoskeleton. Moreover, in the cisternal organelle of cultured hippocampal neurons, ?-actinin colocalizes extensively with synaptopodin, a protein that interacts with both actin and ?-actinin, and they appear concomitantly during the development of these neurons. Together, these results indicate that ?-actinin and the actin cytoskeleton are important components of the cisternal organelle that are probably required to stabilize the AIS.

Choroid plexus epithelial expression of MDR1 P glycoprotein and multidrug resistance-associated protein contribute to the blood–cerebrospinal-fluid drug-permeability barrier

The blood–brain barrier and a blood–cerebrospinal-fluid (CSF) barrier function together to isolate the brain from circulating drugs, toxins, and xenobiotics. The blood–CSF drug-permeability barrier is localized to the epithelium of the choroid plexus (CP). However, the molecular mechanisms regulating drug permeability across the CP epithelium are defined poorly. Herein, we describe a drug-permeability barrier in human and rodent CP mediated by epithelial-specific expression of the MDR1 (multidrug resistance) P glycoprotein (Pgp) and the multidrug resistance-associated protein (MRP). Noninvasive single-photon-emission computed tomography with 99mTc-sestamibi, a membrane-permeant radiopharmaceutical whose transport is mediated by both Pgp and MRP, shows a large blood-to-CSF concentration gradient across intact CP epithelium in humans in vivo. In rats, pharmacokinetic analysis with 99mTc-sestamibi determined the concentration gradient to be greater than 100-fold. In membrane fractions of isolated native CP from rat, mouse, and human, the 170-kDa Pgp and 190-kDa MRP are identified readily. Furthermore, the murine proteins are absent in CP isolated from their respective mdr1a/1b(−/−) and mrp(−/−) gene knockout littermates. As determined by immunohistochemical and drug-transport analysis of native CP and polarized epithelial cell cultures derived from neonatal rat CP, Pgp localizes subapically, conferring an apical-to-basal transepithelial permeation barrier to radiolabeled drugs. Conversely, MRP localizes basolaterally, conferring an opposing basal-to-apical drug-permeation barrier. Together, these transporters may coordinate secretion and reabsorption of natural product substrates and therapeutic drugs, including chemotherapeutic agents, antipsychotics, and HIV protease inhibitors, into and out of the central nervous system.

Dehydroepiandrosterone: A potential signalling molecule for neocortical organization during development

Dehydroepiandrosterone (DHEA) and its sulfate derivative (DHEAS) are the most abundant steroids produced by the human adrenal, but no receptors have been identified for these steroids, and no function for them has been established, other than as precursors for sex steroid synthesis. DHEA and DHEAS are found in brains from many species, and we have shown that enzymes crucial for their synthesis, especially P450c17 (17α-hydroxylase/c17,20 lyase), are expressed in a developmentally regulated, region-specific fashion in the developing rodent brain. One region of embryonic expression of P450c17, the neocortical subplate, has been postulated to play a role in guiding cortical projections to their appropriate targets. We therefore determined if products of P450c17 activity, DHEA and DHEAS, regulated the motility and/or growth of neocortical neurons. In primary cultures of mouse embryonic neocortical neurons, DHEA increased the length of neurites containing the axonal marker Tau-1, and the incidence of varicosities and basket-like process formations in a dose-dependent fashion. These effects could be seen at concentrations normally found in the brain. By contrast, DHEAS had no effect on Tau-1 axonal neurites but increased the length of neurites containing the dendritic marker microtubule-associated protein-2. DHEA rapidly increased free intracellular calcium via activation of N-methyl-d-aspartate (NMDA) receptors. These studies provide evidence of mechanisms by which DHEA and DHEAS exert biological actions, show that they have specific functions other than as sex steroid precursors, mediate their effects via non-classic steroid hormone receptors, and suggest that their developmentally regulated synthesis in vivo may play crucial and different roles in organizing the neocortex.