Yu Ping Tang

Enhanced glial cell line-derived neurotrophic factor mRNA expression upon (−)-deprenyl and melatonin treatments

Glial cell line‐derived neurotrophic factor (GDNF) has been shown to be a preferentially selective neurotrophic factor for dopamine (DA) neurons. In the present study, we have examined the distribution of GDNF mRNA expression in several major DA‐containing cell body and terminal areas and the regulation of GDNF mRNA expression upon various pharmacological treatments. Results indicated that there is a relatively higher GDNF mRNA level in neurons of the nigrostriatal and mesolimbic dopaminergic pathways. Upon chronic 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) treatment (30 mg/kg, i.p., for 7 days), DA level was decreased, whereas GDNF mRNA expression was increased in the striatum, suggesting that more GDNF is synthesized and expressed to cope with the neurotoxin insult. Furthermore, among several DA neuron protective and/or therapeutic agents examined, both intrastriatal injections of (−)‐deprenyl (1.25 μg and 2.5 μg) and melatonin (30 μg, 60 μg, and 120 μg) significantly enhanced GDNF mRNA expression in the striatum, whereas the same concentrations of (−)‐deprenyl did not affect monoamine oxidase B (MAOB) activity, although it increased glutathione peroxidase (GPx) and/or superoxide dismutase (SOD) activities. Similarly, the same concentrations of melatonin did not alter SOD or GPx activities, except that the highest dose of melatonin (120 μg) increased lipid peroxidation in the striatum. Conversely, GM1 ganglioside injection (45 μg) lacked of an effect on GDNF mRNA expression. Together, these results suggest that both (−)‐deprenyl and melatonin up‐regulate GDNF gene expression at threshold doses lower than that needed for altering MAOB activity and/or the antioxidant enzyme systems, respectively. These results provide new information on the neuroprotective and therapeutic mechanisms of (−)‐deprenyl and melatonin on DA neurons. J. Neurosci. Res. 53:593–604, 1998.

Estrogen increases brain expression of the mRNA encoding transthyretin, an amyloid β scavenger protein

Estrogen replacement therapy in postmenopausal women is associated with a reduced risk of Alzheimers Disease (AD). The multiple mechanisms by which estrogen protects against AD are still unknown. To conduct a broad screen for estrogen-regulated AD-related genes in the brain, we used cDNA array assays of brain mRNA samples from ovariectomized (ovx) adult female mice treated with either 17beta-estradiol or vehicle at 1 or 5 weeks post-ovx. The gene encoding transthyretin (TTR), which has been reported to scavenge amyloid beta peptides and reduce amyloid plaque formation, is increased by estradiol treatment at both 1 and 5 weeks post-ovx. Northern blot analyses and RNase protection assays performed on whole brain samples obtained from estradiol- or vehicle-treated mice confirmed the cDNA array assays showing a significant increase in TTR mRNA with estradiol treatment. Qualitative in situ hybridization or immunocytochemistry performed on brain sections demonstrated that TTR mRNA is expressed only in choroid plexus and leptomeninges, and that both estrogen receptor proteins, alpha and beta, are present in choroid plexus cells. These novel findings suggest that estrogen may reduce the risk of AD by acting on choroid plexus cells to increase TTR gene expression, leading to enhanced sequestration and reduced aggregation of amyloid beta peptides.

Stress and corticotropin-releasing factor potentiate center region activity of mice in an open field

The effects of corticotropin-releasing factor (CRF) and previously published effect of stress on the locomotor activity of mice in different regions of an open field were compared. Intracerebroventricular (ICV) administration of 0.2 μg CRF, like stress, significantly increased center region activity; this effect was reversed by the benzodiazepine diazepam (DZP) at a dose of DZP having no significant effect alone. A dose of DZP that antagonized CRF-potentiated center region activity did not block amphetamine-stimulated center area activity. These results suggest that CRF may normally be responsible for many behavioral changes during conditions of stress.

17β-Estradiol Regulates the Sexually Dimorphic Expression of BDNF and TrkB Proteins in the Song System of Juvenile Zebra Finches

Mature brain derived neurotrophic factor (BDNF) plays critical roles in development of brain structure and function, including neurogenesis, axon growth, cell survival and processes associated with learning. Expression of this peptide is regulated by estradiol (E2). The zebra finch song system is sexually dimorphic – only males sing and the brain regions controlling song are larger and have more cells in males compared to females. Masculinization of this system is partially mediated by E2, and earlier work suggests that BDNF with its high affinity receptor TrkB may also influence this development. The present study evaluated expression of multiple forms of both BDNF and TrkB in the developing song system in juvenile males and females treated with E2 or a vehicle control. Using immunohistochemistry and Western blot analysis, BDNF was detected across the song nuclei of 25-day-old birds. Westerns allowed the pro- and mature forms of BDNF to be individually identified, and proBDNF to be quantified. Several statistically significant effects of sex existed in both the estimated total number of BDNF+ cells and relative concentration of proBDNF, varying across the regions and methodologies. E2 modulated BDNF expression, although the specific nature of the regulation depended on brain region, sex and the technique used. Similarly, TrkB (both truncated and full-length isoforms) was detected by Western blot in the song system of juveniles of both sexes, and expression was regulated by E2. In the context of earlier research on these molecules in the developing song system, this work provides a critical step in describing specific forms of BDNF and TrkB, and how they can be mediated by sex and E2. As individual isoforms of each can have opposing effects on mechanisms, such as cell survival, it will now be important to investigate in depth their specific functions in song system maturation.

Gabaergic interneurons in the dorsal raphe mediate the effects of apomorphine on serotonergic system

Apomorphine (APO) has been shown to elevate the concentrations of serotonin (5-HT) and its major metabolite 5-hydroxyindoleacetic acid (5-HIAA) in the mesostriatal but not the mesolimbic serotonergic systems. We have previously demonstrated that the serotonergic actions of APO were secondary to dopamine (DA) autoreceptor stimulation in the substantia nigra. Using picrotoxin as a pharmacological tool, we have presently found that these effects of APO were also indirectly mediated through gamma-aminobutyric acid (GABA) neurons. In examination of the exact anatomical locus of GABA neurons responsible for the observed effects of APO, the results indicate that bilateral lateral habenular lesions did not block the effects of APO on 5-HT neurons, while direct picrotoxin infusion to the dorsal raphe, at a dose having no significant influence by itself, antagonized APOs actions. Together with the anatomical, biochemical and histofluorescent findings, it is suggested that APO influences dorsal raphe 5-HT by stimulation of DA autoreceptors in the substantia nigra; therefore, inhibition of DA neuron activity and the nigro-raphe pathway. Normally, DA probably exerts an excitatory influence on gabaergic interneurons in the dorsal raphe, and these inhibitory interneurons then synapse on 5-HT neurons in the same area. Activation of 5-HT neurons were explained by a disinhibitory effect as a result of reduced release of GABA due to feedback inhibition of DA neuron firing following APO activation of DA autoreceptors in the substantia nigra. The striatal presynaptic and postsynaptic DA receptors, however, do not appear to mediate the above effects of APO.

Sexually dimorphic expression of the genes encoding ribosomal proteins L17 and L37 in the song control nuclei of juvenile zebra finches

Studies evaluating the role of steroid hormones in sexual differentiation of the zebra finch song system have produced complicated and at times paradoxical results, and indicate that additional factors may be critical. Therefore, in a previous study we initiated a screen for differential gene expression in the telencephalon of developing male and female zebra finches. The use of cDNA microarrays and real-time quantitative PCR revealed increased expression of the genes encoding ribosomal proteins L17 and L37 (RPL17 and RPL37) in the male forebrain as a whole. Preliminary in situ hybridization data then indicated enhanced expression of both these genes in song control regions. Two experiments in the present study quantified the mRNA expression. The first utilized 25-day-old male and female zebra finches. The second compared a separate set of juveniles to adults of both sexes to both re-confirm enhanced expression in juvenile males and to determine whether it is limited to developing animals. In Experiment 1, males exhibited increased expression of both RPL17 and RPL37 compared to females in Area X, the robust nucleus of the arcopallium (RA), and the ventral ventricular zone (VVZ), which may provide neurons to Area X. Experiment 2 replicated the sexually dimorphic expression of these genes at post-hatching day 25, and documented that the sex differences are eliminated or greatly reduced in adults. The results are consistent with the idea that these ribosomal proteins may influence sexual differentiation of Area X and RA, potentially regulating the genesis and/or survival of neurons.

Photoperiod Modulates Pubertal Shifts in Behavioral Responsiveness to Testosterone

This study examined the effect of photoperiod on pubertal maturation of steroid-dependent reproductive behaviors in male European ferrets (Mustela putoriusfuro). In the first experiment, levels of neck gripping, mounting, and pelvic thrusting in gonadally intact prepubertal (PRE) ferrets were compared with those of adults that had undergone puberty either while housed in short days (8 hr light/16 hr darkness per day; SD), or after transfer from SD to long days (18 hr light/6 hr darkness per day; LD) at 12 weeks of age. Both LD and SD adults demonstrated significantly greater amounts of neck gripping and mounting than PRE males. In addition, a significantly greater proportion of adults in both SD and LD displayed at least one incidence of the three behaviors compared to PRE ferrets. There were no statistically significant differences in behavior of the gonadally intact LD and SD adults. In the second experiment, dose-response curves for behavioral responses to subcutaneous injections of 0, 0.5, 1.25, 2.5, 5, and 10 mg/kg testosterone propionate (TP) in oil were generated in castrated PRE, SD, and LD males. The lowest dose of TP elicited significantly greater amounts of all three behaviors in LD adults than in PRE ferrets. In addition, levels of mounting and thrusting elicited by the lowest dose of TP were significantly greater in LD adults than in SD adults. These data indicate that pubertal activation of male sexual behavior in male ferrets is accompanied by a pubertal increase in responsiveness to the behavioral effects of testosterone. Furthermore, the degree of behavioral responsiveness of adult ferrets to testosterone is modulated by environmental photoperiod experienced during reproductive maturation.

Effects of Estradiol on Incorporation of New Cells in the Developing Zebra Finch Song System: Potential Relationship to Expression of Ribosomal Proteins L17 and L37

Mechanisms regulating masculinization of the zebra finch song system are unclear; both estradiol and sex‐specific genes may be important. This study was designed to investigate relationships between estrogen and ribosomal proteins (RPL17 and RPL37; sex‐linked genes) that exhibit greater expression in song control nuclei in juvenile males than females. Four studies on zebra finches were conducted using bromodeoxyuridine (BrdU) injections on posthatching days 6–10 with immunohistochemistry for the ribosomal proteins and the neuronal marker HuC/D at day 25. Volumes of brain regions were also assessed in Nissl‐stained tissue. Most BrdU+ cells expressed RPL17 and RPL37. The density and percentage of cells co‐expressing BrdU and HuC/D was greatest in Area X. The density of BrdU+ cells in Area X (or its equivalent) and the percentage of these cells that were neurons were greater in males than females. In RA and HVC, total BrdU+ cells were increased in males. A variety of effects of estradiol were also detected, including inducing an Area X in females with a masculine total number of BrdU+ cells, and increasing the volume and percentage of new neurons in the HVC of females. The same manipulation in males decreased the density of BrdU+ cells in Area X, total number of BrdU+ cells in RA, and density of new neurons in HVC and RA. These data are consistent with the idea that RPL17, RPL37, and estradiol might all influence sexual differentiation, perhaps with the hormone and proteins interacting, such that an appropriate balance is required for normal development.

Differential biochemical mechanisms mediate locomotor stimulation effects by caffeine and nicotine in rats.

Effects of caffeine and the interactive effects of caffeine and nicotine on locomotor activity in rats were examined in the present study. Other than confirming previous reports that both drugs enhanced locomotion, we have also found that their effects on activity were additive. Meanwhile, results of various biochemical measures have revealed that at the minimum effective doses of caffeine and nicotine which facilitated locomotor activity, only one biochemical system was preferentially influenced by either drug alone. The most significant findings were that caffeine stimulated the release of catecholamines and nicotine decreased the concentrations of tyrosine and tryptophan in brain. The combined effects of caffeine and nicotine on these brain amines were not different from those of each drug alone. Together with the report that caffeine and nicotine had differential actions on different activity measures, the present results support the hypothesis that caffeine and nicotine affect locomotor activity via different neurochemical mechanisms.

Co-localization of Sorting Nexin 2 and androgen receptor in the song system of juvenile zebra finches

Mechanisms regulating sexual differentiation of the zebra finch song system appear to include both genetic and hormonal factors. Sorting Nexin 2 (SNX2), which is involved in trafficking proteins between cellular membranes, and androgen receptor (AR) mRNA are both increased in song control nuclei of juvenile males compared to females. Here, in situ hybridization for SNX2 and immunohistochemistry for AR were used to evaluate these sexual dimorphisms in more detail. Estimates of the total number of HVC cells expressing SNX2 and AR, individually as well as together, were greater in 25-day-old males compared to females. The densities of these types of cells were generally also increased in males compared to females in HVC and Area X (or the equivalent portion of the medial striatum in females). On average, more than half of the AR+ cells co-expressed SNX2 in both brain regions. The potential, therefore, exists for both AR and SNX2 to be involved in masculinization of these two brain regions. One possibility is that they, either separately or in conjunction, enhance the action of trophic factors within the brain.