SOX2 is sequentially required for progenitor proliferation and lineage specification in the developing pituitary.

Sox2 mutations are associated with pituitary hormone deficiencies and the protein is required for pituitary progenitor proliferation, but its function has not been well characterized in this context. SOX2 is known to activate expression of Six6, encoding a homeodomain transcription factor, in the ventral diencephalon. Here, we find that the same relationship likely exists in the pituitary. Moreover, because Six6 deletion is associated with a similar phenotype as described here for loss of Sox2, Six6 appears to be an essential downstream target of SOX2 in the gland. We also uncover a second role for SOX2. Whereas cell differentiation is reduced in Sox2 mutants, some endocrine cells are generated, such as POMC-positive cells in the intermediate lobe. However, loss of SOX2 here results in complete downregulation of the melanotroph pioneer factor PAX7, and subsequently a switch of identity from melanotrophs to ectopic corticotrophs. Rescuing proliferation by ablating the cell cycle negative regulator p27 (also known as Cdkn1b) in Sox2 mutants does not restore melanotroph emergence. Therefore, SOX2 has two independent roles during pituitary morphogenesis; firstly, promotion of progenitor proliferation, and subsequently, acquisition of melanotroph identity.

Illumination controls differentiation of dopamine neurons regulating behaviour.

Specification of the appropriate neurotransmitter is a crucial step in neuronal differentiation because it enables signalling among populations of neurons. Experimental manipulations demonstrate that both autonomous and activity-dependent genetic programs contribute to this process during development, but whether natural environmental stimuli specify transmitter expression in a neuronal population is unknown. We investigated neurons of the ventral suprachiasmatic nucleus that regulate neuroendocrine pituitary function in response to light in teleosts, amphibia and primates. Here we show that altering light exposure, which changes the sensory input to the circuit controlling adaptation of skin pigmentation to background, changes the number of neurons expressing dopamine in larvae of the amphibian Xenopus laevis in a circuit-specific and activity-dependent manner. Neurons newly expressing dopamine then regulate changes in camouflage colouration in response to illumination. Thus, physiological activity alters the numbers of behaviourally relevant amine-transmitter-expressing neurons in the brain at postembryonic stages of development. The results may be pertinent to changes in cognitive states that are regulated by biogenic amines.

Pituitary pars intermedia dysfunction with pituitary gland melanotroph adenoma in a Grevy's zebra (Equus grevyi).

A 22-year and 9-month-old female Grevy's zebra (Equus grevyi) showed signs of polyuria, polydipsia, glucosuria, and muscle atrophy. Blood tests revealed hyperglycemia, hypertriglyceridemia, electrolyte imbalance, high levels of adrenocorticotropic hormone (ACTH) and cortisol, and low levels of hormones secreted by the pituitary pars distalis. Pathological examinations revealed a pituitary gland tumor and bilateral adrenal cortical hyperplasia. Pituitary tumor cells showed immunoreactivity for α-melanocyte-stimulating hormone and ACTH. The deposition of amyloid ß was observed in the parenchyma and vascular walls of the cerebrum. The zebra showed clinical signs of pituitary pars intermedia dysfunction and was histopathologically diagnosed with pituitary gland melanotroph adenoma. This case report provides insight into neoplastic and endocrine diseases associated with the aging of a zebra.

Cytologic features of canine melanotroph and corticotroph pituitary adenomas.

BACKGROUND: The introduction of intraoperative cytology revolutionized neurosurgical procedures in human medicine, providing real-time diagnostic guidance to surgeons and contributing to improved patient outcomes. In the realm of veterinary medicine, the understanding of pituitary tumors in dogs and cats remains limited due to challenges in obtaining antemortem samples of central nervous system lesions. OBJECTIVES: The aim of this study was to describe the cytologic features of pituitary adenomas in 12 dogs that underwent hypophysectomy. METHODS: The series included nine melanotroph adenomas and three corticotroph adenomas. Definitive diagnosis was based on histopathology and immunohistochemistry. RESULTS: Cytologically, the adenomas had high numbers of bare nuclei and intact cells that were round to polygonal and situated individually or in small clusters. The intact cells had round to oval, eccentric nuclei with finely stippled chromatin and one to three prominent nucleoli and ample to abundant lightly basophilic to amphophilic, grainy cytoplasm with distinct borders, and variable numbers of discrete vacuoles. Mild-to-moderate anisocytosis and anisokaryosis, occasional binucleation, rare and atypical mitotic figures, and nuclear molding were also observed. CONCLUSIONS: The results suggest that intraoperative cytology of canine pituitary adenomas holds promise as a valuable diagnostic tool, aiding swift differentiation from other sellar masses before histologic confirmation. Cytologic characterization of pituitary adenomas in dogs is exceptionally rare in the scientific literature, making this study one of the first to offer a comprehensive description of these cytologic features.

Protein Tyrosine Phosphatase Receptors N and N2 Control Pituitary Melanotroph Development and POMC Expression.

The neuroendocrine marker genes Ptprn and Ptprn2 encode protein tyrosine phosphatase receptors N and N2, 2 members of protein tyrosine phosphatase receptors void of enzymatic activity, and whose function and mechanism of action have not been elucidated. To explore the role(s) of Ptprn and Ptprn2 on the hypothalamic-pituitary-adrenal axis, we used mice in which both genes were knocked out (DKO). The focus in this study was on corticotrophs and melanotrophs from the anterior and intermediate lobes of the pituitary gland, respectively. In both sexes, DKO caused an increase in the expression of the corticotroph/melanotroph genes Pomc and Tbx19 and the melanotroph-specific gene Pax7. We also found in vivo and in vitro increased synthesis and release of beta-endorphin, alpha-melanocyte-stimulating hormone, and ACTH in DKO mice, which was associated with increased serum corticosterone levels and adrenal mass. DKO also increased the expression of other melanotroph-specific genes, but not corticotroph-specific genes. The dopaminergic pathway in the hypothalamus and dopaminergic receptors in melanotrophs were not affected in DKO mice. However, hyperplasia of the intermediate lobe was observed in DKO females and males, accompanied by increased proopiomelanocortin immunoreactivity per cell. These results indicate that protein tyrosine phosphatase receptor type N contributes to hypothalamic-pituitary-adrenal function by being involved in processes governing postnatal melanotroph development and Pomc expression.

Rab3a ablation related changes in morphology of secretory vesicles in major endocrine pancreatic cells, pituitary melanotroph cells and adrenal gland chromaffin cells in mice.

In this work we have compared the ultrastructural characteristics of major pancreatic endocrine cells, pituitary melanotrophs and adrenal chromaffin cells in the normal mouse strain (wild type, WT) and mice with a known secretory deficit, the Rab3a knockout strain (Rab3a KO). For this purpose, pancreata, pituitary glands and adrenal glands from the Rab3a KO and from the WT mice were analysed, using conventional transmission electron microscopy (TEM). In order to assess the significance of the presence of Rab3a proteins in the relevant cells, we focused primarily on their secretory vesicle morphology and distribution. Our results showed a comparable general morphology in Rab3a KO and WT in all assessed endocrine cell types. In all studied cell types, the distribution of secretory granules along the plasma membrane (number of docked and almost-docked vesicles) was comparable between Rab3a KO and WT mice. Specific differences were found in the diameters of their secretory vesicles, diameters of their electron-dense cores and the presence of autophagic structures in the cells of Rab3A KO mice only. Occasionally, individual electron-dense round vesicles were present inside autophagosome-like structures; these were possibly secretory vesicles or their remnants. The differences found in the diameters of the secretory vesicles confirm the key role of Rab3a proteins in controlling the balance between secretory vesicle biogenesis and degradation, and suggest that the ablation of this protein probably changes the nature of the reservoir of secretory vesicles available for regulated exocytosis.

Persistent expression of activated Notch inhibits corticotrope and melanotrope differentiation and results in dysfunction of the HPA axis.

The hypothalamic-pituitary-adrenal (HPA) axis is an important regulator of energy balance, immune function and the body's response to stress. Signaling networks governing the initial specification of corticotropes, a major component of this axis, are not fully understood. Loss of function studies indicate that Notch signaling may be necessary to repress premature differentiation of corticotropes and to promote proliferation of pituitary progenitors. To elucidate whether Notch signaling must be suppressed in order for corticotrope differentiation to proceed and whether Notch signaling is sufficient to promote corticotrope proliferation, we examined the effects of persistent Notch expression in Pomc lineage cells. We show that constitutive activation of the Notch cascade inhibits the differentiation of both corticotropes and melanotropes and results in the suppression of transcription factors required for Pomc expression. Furthermore, persistent Notch signaling traps cells in the intermediate lobe of the pituitary in a progenitor state, but has no effect on pituitary proliferation. Undifferentiated cells are eliminated in the first two postnatal weeks in these mice, resulting in a modest increase in CRH expression in the paraventricular nucleus, hypoplastic adrenal glands and decreased stress-induced corticosterone levels. Taken together, these findings show that Notch signaling is sufficient to prevent corticotrope and melanotrope differentiation, resulting in dysregulation of the HPA axis.

Gene expression profiling of pituitary melanotrope cells during their physiological activation.

The pituitary melanotrope cells of the amphibian Xenopus laevis are responsible for the production of the pigment-dispersing peptide α-melanophore-stimulating hormone, which allows the animal to adapt its skin color to its environment. During adaptation to a dark background the melanotrope cells undergo remarkable changes characterized by dramatic increases in cell size and secretory activity. In this study we performed microarray mRNA expression profiling to identify genes important to melanotrope activation and growth. We show a strong increase in the expression of the immediate early gene (IEG) c-Fos and of the brain-derived neurotrophic factor gene (BDNF). Furthermore, we demonstrate the involvement of another IEG in the adaptation process, Nur77, and conclude from in vitro experiments that the expression of both c-Fos and Nur77 are partially regulated by the adenylyl cyclase system and calcium ions. In addition, we found a steady up-regulation of Ras-like product during the adaptation process, possibly evoked by BDNF/TrkB signaling. Finally, the gene encoding the 105-kDa heat shock protein HSPh1 was transiently up-regulated in the course of black-background adaptation and a gene product homologous to ferritin (ferritin-like product) was >100-fold up-regulated in fully black-adapted animals. We suggest that these latter two genes are induced in response to cellular stress and that they may be involved in changing the mode of mRNA translation required to meet the increased demand for de novo protein synthesis. Together, our results show that microarray analysis is a valuable approach to identify the genes responsible for generating coordinated responses in physiologically activated cells.

The role of brain-derived neurotrophic factor in the regulation of cell growth and gene expression in melanotrope cells of Xenopus laevis.

Brain-derived neurotrophic factor (BDNF) is, despite its name, also found outside the central nervous system (CNS), but the functional significance of this observation is largely unknown. This review concerns the expression of BDNF in the pituitary gland. While the presence of the neurotrophin in the mammalian pituitary gland is well documented its functional significance remains obscure. Studies on the pars intermedia of the pituitary of the amphibian Xenopus laevis have shown that BDNF is produced by the neuroendocrine melanotrope cells, its expression is physiologically regulated, and the melanotrope cells themselves express receptors for the neurotrophin. The neurotrophin has been shown to act as an autocrine factor on the melanotrope to promote cell growth and regulate gene expression. In doing so BDNF supports the physiological function of the cell to produce and release α-melanophore-stimulating hormone for the purpose of adjusting the animal's skin color to that of its background.

Pituitary melanotrope cells of Xenopus laevis are of neural ridge origin and do not require induction by the infundibulum.

Classical studies in amphibians have concluded that the endocrine pituitary and pars intermedia are derived from epithelial buccal epidermis and do not require the infundibulum for their induction. These studies also assumed that the pituitary is not subsequently determined by infundibular induction. Our extirpation, auto-transplantation and immunohistochemical studies with Xenopus laevis were initiated to investigate early presumptive pituitary development. These studies were conducted especially with reference to the pars intermedia melanotrope cell's induction, and its production and release of α-melanophore stimulating hormone (α-MSH) from the precursor protein proopiomelanocortin (POMC). Auto-transplantation studies demonstrated that the pituitary POMC-producing cells are determined at a stage prior to pituitary-infundibular contact. The results of experiments involving the extirpation of the presumptive infundibulum also indicated that the infundibulum is not essential for the differentiation of POMC-producing cells. We also demonstrated that early pituitary development involves adherence to the prechiasmatic area of the diencephalon with the pituitary placode growing in a posterior direction toward the infundibulum where contact occurs at Xenopus stage 39/40. Overall, our studies provide a model for early tissue relations among presumptive pituitary, suprachiasmatic nucleus, pars tuberalis and infundibulum during neurulation and later neural tube stages of development. It is hypothesized that the overlying chiasmatic area suppresses pituitary differentiation.

Continuous stress-induced dopamine dysregulation augments PAP-I and PAP-II expression in melanotrophs of the pituitary gland.

Under continuous stress (CS) in rats, melanotrophs, the predominant cell-type in the intermediate lobe (IL) of the pituitary, are hyperactivated to secrete α-melanocyte-stimulating hormone and thereafter degenerate. Although these phenomena are drastic, the molecular mechanisms underlying the cellular changes are mostly unknown. In this study, we focused on the pancreatitis-associated protein (PAP) family members of the secretory lectins and characterized their expression in the IL of CS model rats because we had identified two members of this family as up-regulated genes in our previous microarray analysis. RT-PCR and histological studies demonstrated that prominent PAP-I and PAP-II expression was induced in melanotrophs in the early stages of CS, while another family member, PAP-III, was not expressed. We further examined the regulatory mechanisms of PAP-I and PAP-II expression and revealed that both were induced by the decreased dopamine levels in the IL under CS. Because the PAP family members are implicated in cell survival and proliferation, PAP-I and PAP-II secreted from melanotrophs may function to sustain homeostasis of the IL under CS conditions in an autocrine or a paracrine manner.

Analysis of the melanotrope cell neuroendocrine interface in two amphibian species, Rana ridibunda and Xenopus laevis: a celebration of 35 years of collaborative research.

This review gives an overview of the functioning of the hypothalamo-hypophyseal neuroendocrine interface in the pituitary neurointermediate lobe, as it relates to melanotrope cell function in two amphibian species, Rana ridibunda and Xenopus laevis. It primarily but not exclusively concerns the work of two collaborating laboratories, the Laboratory for Molecular and Cellular Neuroendocrinology (University of Rouen, France) and the Department of Cellular Animal Physiology (Radboud University Nijmegen, The Netherlands). In the course of this review it will become apparent that Rana and Xenopus have, for the most part, developed the same or similar strategies to regulate the release of α-melanophore-stimulating hormone (α-MSH). The review concludes by highlighting the molecular and cellular mechanisms utilized by thyrotropin-releasing hormone (TRH) to activate Rana melanotrope cells and the function of autocrine brain-derived neurotrophic factor (BDNF) in the regulation of Xenopus melanotrope cell function.

Brain-derived neurotrophic factor stimulates growth of pituitary melanotrope cells in an autocrine way.

Brain-derived neurotrophic factor (BDNF) is expressed in the mammalian pituitary gland, in both the anterior and intermediate lobes, where its functional significance is unknown. Melanotrope cells in the intermediate pituitary lobe of the amphibian Xenopus laevis also produce BDNF, which co-exists in secretory granules with α-melanophore-stimulating hormone (α-MSH), a peptide that causes pigment dispersion in dermal melanophores during adaptation of the toad to a dark background. Xenopus melanotropes are highly plastic, undergoing very strong growth to support the high biosynthesis and release of α-MSH in black-adapted animals. In this study we have tested our hypothesis that this enhanced growth of the melanotrope is maintained by autocrine release of BDNF. Furthermore, since the extracellular-regulated kinase (ERK) pathway is a major component of BDNF signaling in neuronal plasticity, we investigated its involvement in melanotrope cell growth. For these purposes melanotropes were treated for 3 days in vitro, with either an anti-BDNF serum or a recombinant tropomyosin-receptor kinase B (TrkB) receptor fragment to eliminate released BDNF, or with the ERK inhibitor U0126. We also applied a novel inhibitor of the TrkB receptor, cyclotraxin-B, to test this receptor's involvement in melanotrope cell growth regulation. All treatments markedly reduced melanotrope cell growth. Therefore, we conclude that autocrine release of BDNF and subsequent TrkB-dependent ERK-mediated signaling is important for melanotrope cell growth during its physiologically induced activation.

Expression patterns of hormones, signaling molecules, and transcription factors during adenohypophysis development in the chick embryo.

The chick embryo is an ideal model to study pituitary cell-type differentiation. Previous studies describing the temporal appearance of differentiated pituitary cell types in the chick embryo are contradictory. To resolve these controversies, we used RT-PCR to define the temporal onset and in situ hybridization and immunohistochemistry to define the spatial localization of hormone expression within the pituitary. RT-PCR detected low levels of Fshbeta (gonadotropes) and Pomc (corticotropes, melanotropes) mRNA at E4 and Gh (somatotropes), Prl (lactotropes), and Tshbeta (thyrotropes) mRNA at E8. For all hormones, sufficient accumulation of mRNA and/or protein to permit detection by in situ hybridization or immunohistochemistry was observed approximately 3 days later and in all cases corresponded to a notable increase in RT-PCR product. We also describe the expression patterns of signaling (Bmp2, Bmp4, Fgf8, Fgf10, Shh) and transcription factors (Pitx1, Pitx2, cLim3) known to be important for pituitary organogenesis in other model organisms.

Activating mutations in BRAF disrupt the hypothalamo-pituitary axis leading to hypopituitarism in mice and humans.

Germline mutations in BRAF and other components of the MAPK pathway are associated with the congenital syndromes collectively known as RASopathies. Here, we report the association of Septo-Optic Dysplasia (SOD) including hypopituitarism and Cardio-Facio-Cutaneous (CFC) syndrome in patients harbouring mutations in BRAF. Phosphoproteomic analyses demonstrate that these genetic variants are gain-of-function mutations leading to activation of the MAPK pathway. Activation of the MAPK pathway by conditional expression of the BrafV600E/+ allele, or the knock-in BrafQ241R/+ allele (corresponding to the most frequent human CFC-causing mutation, BRAF p.Q257R), leads to abnormal cell lineage determination and terminal differentiation of hormone-producing cells, causing hypopituitarism. Expression of the BrafV600E/+ allele in embryonic pituitary progenitors leads to an increased expression of cell cycle inhibitors, cell growth arrest and apoptosis, but not tumour formation. Our findings show a critical role of BRAF in hypothalamo-pituitary-axis development both in mouse and human and implicate mutations found in RASopathies as a cause of endocrine deficiencies in humans.

Expression and Role of Thyrotropin Receptors in Proopiomelanocortin-Producing Pituitary Cells.

Background: Thyrotropin (TSH) is well known as the hormone of the anterior pituitary thyrotrophs responsible for acting in the thyroid gland, where it stimulates synthesis and release of thyroid hormones through Gs and Gq/11 protein coupled TSH receptors (TSHRs). Methods: In this study, we examined whether the functional TSHRs are also expressed in cultured rat pituitary cells, using double immunocytochemistry, quantitative reverse transcription-polymerase chain reaction analysis, cAMP and hormone measurements, and single-cell calcium imaging. Results: Double immunocytochemistry revealed the expression of TSHRs in cultured corticotrophs and melanotrophs, in addition to previously identified receptors in folliculostellate cells. The functional coupling of these receptors to the Gq/11 signaling pathway was not observed, as demonstrated by the lack of TSH activation of IP3-dependent calcium mobilization in these cells when bathed in calcium-deficient medium. However, TSH increased cAMP production in a time- and concentration-dependent manner and facilitated calcium influx in single corticotrophs and melanotrophs, indicating their coupling to the Gs signaling pathway. Consistent with these findings, TSH stimulated adrenocorticotropin and ß-endorphin release in male and female pituitary cells in a time- and concentration-dependent manner without affecting the expression of proopiomelanocortin gene. Conclusions: These results indicate that TSH is a potential paracrine modulator of anterior pituitary corticotrophs and melanotrophs, controlling the exocytotic but not the transcriptional pathway in a cAMP/calcium influx-dependent manner.

A proteome map of the pituitary melanotrope cell activated by black-background adaptation of Xenopus laevis.

Upon transfer of Xenopus laevis from a white to a black background, the melanotrope cells in the pituitary pars intermedia secrete alpha-melanocyte-stimulating hormone, which stimulates dispersion of melanin pigment in skin melanophores. This adaptive behavior is under the control of neurotransmitters and neuropeptides of hypothalamic origin. The alpha-melanocyte-stimulating hormone-producing cells and their hypothalamic control system provide an interesting model to study proteins required for biosynthetic and secretory processes involved in peptide hormone production and for brain-pituitary signaling. We present a 2-D PAGE-based proteome map of melanotrope cells from black-adapted animals, identifying 204 different proteins by MS analysis.

Plasticity of melanotrope cell regulations in Xenopus laevis.

This review focuses on the plasticity of the regulation of a particular neuroendocrine transducer cell, the melanotrope cell in the pituitary pars intermedia of the amphibian Xenopus laevis. This cell type is a suitable model to study the relationship between various external regulatory inputs and the secretion of an adaptive endocrine message, in this case the release of α-melanophore-stimulating hormone, which activates skin melanophores to darken when the animal is placed on a dark background. Information about the environmental conditions is processed by various brain centres, in the hypothalamus and elsewhere, that eventually control the activity of the melanotrope cell regarding hormone production and secretion. The review discusses the roles of these hypothalamic and extrahypothalamic nuclei, their neurochemical messengers acting on the melanotrope, and the external stimuli they mediate to control melanotrope cell functioning.

Functional diversity among p24 subfamily members.

BACKGROUND INFORMATION: The p24 protein family plays an important but unclear role at the ER (endoplasmic reticulum)-Golgi interface. A p24 member from each subfamily (p24alpha(3), beta(1), gamma(3) and delta(2)) is upregulated with the prohormone POMC (pro-opiomelanocortin) when Xenopus laevis intermediate pituitary melanotrope cells are physiologically activated. Here we explored the role of p24 by generating and analysing Xenopus with melanotrope cell-specific transgene expression of p24beta(1) or p24gamma(3), two of the p24 proteins coexpressed with POMC, and compared the results with those previously reported for the two other coexpressed p24s (p24alpha(3) and p24delta(2)). RESULTS: The transgene expression of p24beta(1) or p24gamma(3) did not affect the endogenous p24 proteins or affected only endogenous p24gamma(3) respectively, whereas in transgenics expressing p24alpha(3) and p24delta(2), the levels of all endogenous p24 proteins were strongly decreased. Nevertheless, as for p24alpha(3) but albeit to a lesser extent, in the p24beta(1)-transgenic melanotrope cells the rate of cargo cleavage was reduced, probably reflecting reduced cargo transport from the ER, and POMC glycosylation and sulfation in the Golgi were not affected. The p24gamma(3)-transgenic cells displayed features of both the p24alpha(3)-transgenics (reduced cargo cleavage, normal POMC sulfation) and the p24delta(2)-transgenics (affected POMC glycosylation). CONCLUSIONS: Our results show that the four upregulated proteins p24alpha(3), beta(1), gamma(3) and delta(2) have non-redundant roles in the early secretory pathway, and suggest that each p24 subfamily member provides a proper ER/Golgi subcompartmental microenvironment, together allowing correct secretory protein transport and processing.

BDNF stimulates Ca2+ oscillation frequency in melanotrope cells of Xenopus laevis: contribution of IP3-receptor-mediated release of intracellular Ca2+ to gene expression.

Pituitary melanotrope cells of the amphibian Xenopus laevis are neuroendocrine cells regulating the animal's skin color adaptation through secretion of α-melanophore-stimulating hormone (α-MSH). To fulfill this function optimally, the melanotrope cell undergoes plastic changes in structure and secretory activity in response to changed background light conditions. Xenopus melanotrope cells display Ca(2+) oscillations that are thought to drive α-MSH secretion and gene expression. They also produce brain-derived neurotrophic factor (BDNF), which stimulates in an autocrine way the biosynthesis of the α-MSH precursor, pro-opiomelanocortin (POMC). We have used this physiological adaptation mechanism as a model to investigate the role of BDNF in the regulation of Ca(2+) kinetics and Ca(2+)-dependent gene expression. By dynamic video imaging of isolated cultured melanotropes we demonstrated that BDNF caused a dose-dependent increase in Ca(2+) oscillation frequency up to 64.7±2.3% of control level. BDNF also induced a transient Ca(2+) peak in Ca(2+)-free medium, which was absent when calcium stores were blocked by thapsigargin and 2-aminoethoxydiphenyl borate, indicating that BDNF stimulates acute release of Ca(2+) from IP(3)-sensitive intracellular Ca(2+) stores. Moreover, we show that thapsigargin inhibits the expression of BDNF transcript IV (by 61.1±28.8%) but does not affect POMC transcript. We conclude that BDNF mobilizes Ca(2+) from IP(3)-sensitive intracellular Ca(2+) stores and propose the possibility that the resulting Ca(2+) oscillations selectively stimulate expression of the BDNF gene.