Early and late preterm birth rates in participants adherent to randomly assigned high dose docosahexaenoic acid (DHA) supplementation in pregnancy.

BACKGROUND: Intention-to-treat analyses do not address adherence. Per protocol analyses treat nonadherence as a protocol deviation and assess if the intervention is effective if followed. OBJECTIVE: To determine the rate of early preterm birth (EPTB, <34 weeks gestation) and preterm birth (PTB, <37 weeks gestation) in participants who adhered to a randomly assigned docosahexaenoic acid (DHA) dose of 1000 mg/day. STUDY DESIGN: Eleven hundred women with a singleton pregnancy were enrolled before 20-weeks' gestation, provided a capsule with 200 mg/day DHA and randomly assigned to two additional capsules containing a placebo or 800 mg of DHA. In the Bayesian Adaptive Design, new randomization schedules were determined at prespecified intervals. In each randomization, the group with the most EPTB was assigned fewer participants than the other group. Adherence was defined a priori as a postpartum red blood cell phospholipid DHA (RBC-PL-DHA) ≥5.5%.and post hoc as ≥8.0% RBC-PL-DHA, the latter after examination of postpartum RBC-PL-DHA. Bayesian mixture models were fitted for gestational age and dichotomized for EPTB and PTB as a function of baseline RBC-PL-DHA and dose-adherence. Bayesian hierarchical models were also fitted for EPTB by dose adherence and quartiles of baseline RBC-PL-DHA. RESULTS: Adherence to the high dose using both RBC-PL-DHA cut points resulted in less EPTB compared to 200 mg [Bayesian posterior probability (pp) = 0.93 and 0.92, respectively]. For participants in the two lowest quartiles of baseline DHA status, adherence to the higher dose resulted in lower EPTB (≥5.5% RBC-PL-DHA, quartiles 1 and 2, pp = 0.95 and 0.96; ≥8% RBC-PL-DHA, quartiles 1 and 2, pp = 0.94 and 0.95). Using the Bayesian model, EPTB was reduced by 65%, from 3.45% to 1.2%, using both cut points. Adherence also reduced PTB before 35, 36 and 37 weeks using both cut points (pp ≥ 0.95). In general, performance of the nonadherent subgroup mirrored that of participants assigned to 200 mg. CONCLUSION: Adherence to high dose DHA reduced EPTB and PTB. The largest effect of adherence on reducing EPTB was observed in women with low baseline DHA levels. CLINICALTRIALS: gov (NCT02626299).

Validation of an abbreviated food frequency questionnaire for estimating DHA intake of pregnant women in the United States.

Docosahexaenoic acid (DHA) intake was estimated in pregnant women between 12- and 20-weeks' gestation using the National Cancer Institute's (NCI) Diet History Questionnaire-II (DHQ-II) and a 7-question screener designed to capture DHA intake (DHA Food Frequency Questionnaire, DHA-FFQ). Results from both methods were compared to red blood cell phospholipid DHA (RBC-DHA) weight percent of total fatty acids. DHA intake from the DHA-FFQ was more highly correlated with RBC-DHA (rs=0.528) than the DHQ-II (rs=0.352). Moreover, the DHA-FFQ allowed us to obtain reliable intake data from 1355 of 1400 participants. The DHQ-II provided reliable intake for only 847 of 1400, because many participants only partially completed it and it was not validated for Hispanic participants. Maternal age, parity, and socioeconomic status (SES) were also significant predictors of RBC-DHA. When included with estimated intake from the DHA-FFQ, the model accounted for 36% of the variation in RBC-DHA.

Pediatric allo-SCT for malignant and non-malignant diseases: impact on health-related quality of life outcomes.

The objective of this study was prospectively to investigate the health-related quality of life (HRQOL) of 80 pediatric recipients of allo-SCT for malignant and non-malignant diseases. The PedsQL 4.0 was used to assess self-reported physical, emotional and social functioning of children 5 years old once, pre-allo-SCT and on days +100, +180, +365 and +730. Emotional and social functioning was stable pre-to-post-allo-SCT and comparable to the normative sample (P>0.05), and physical functioning was 17 points lower pre-allo-SCT (P0.01) with improved scores equivalent to the norms by day +730. Lower physical scores were reflected by 50-54% of children reporting difficulties with movement, strength, pain and fatigue. At baseline, children ages 5-7 reported lower social functioning (P<0.05) and patients with non-malignant disease reported better physical functioning (P<0.05). Emotional functioning in ages 8-12 improved over time (P<0.05). More than 50% of the participants were minority and their HRQOL was similar to non-minority participants. Physical functioning significantly improved for recipients of reduced-toxicity conditioning (P0.01), significantly worsened for patients with chronic GVHD (cGVHD; P<0.05), and significantly decreased in recipients of matched-unrelated donor transplant who developed cGVHD (P<0.05). Multidisciplinary efforts are necessary to identify and support pediatric patients' physical needs to improve functional outcomes.

Differential regulation of GABA B receptor subunit expression and function.

The GABA(B) receptor is a G protein-coupled heterodimer composed of GABA(B1) and GABA(B2) subunits. In the present study, experiments were undertaken to examine the relationship between GABA(B) receptor function and subunit expression in the rat lumbar spinal cord following pharmacological and physiological manipulation of this receptor system. Although formalin-induced hind paw inflammation increases the production of GABA(B1) and GABA(B2) protein in the spinal cord within 24 h, there is no change in receptor function, as measured by the baclofen-stimulated guanosine 5'-O-(3-[(35)S]thiotriphosphate) ([(35)S]GTPgammaS) binding assay. Conversely, although chronic (7 days) administration of baclofen, a GABA(B) receptor agonist, abolishes baclofen-stimulated [(35)S]GTPgammaS binding in the spinal cord tissue, causes tolerance to the sedative and antinociceptive effects of the drug, increases the number of formalin-induced hind paw flinches, and induces mechanical hyperalgesia, this treatment had no effect on the levels of GABA(B1) or GABA(B2) mRNAs in the lumbar spinal cord. The results indicate a lack of concordance between expression of GABA(B1) and GABA(B2) subunits and GABA(B) receptor function, suggesting these subunit proteins may serve multiple functions in the cells. Moreover, these findings indicate that nongenomic mechanisms are primarily responsible for the GABA(B) receptor desensitization that occurs during prolonged exposure to receptor agonist.

Effects of stress and tranylcypromine on amphetamine-induced locomotor activity and GABA(B) receptor function in rat brain.

Modification in gamma-aminobutyric acid-B (GABA(B)) receptors may contribute to the symptoms of some neurological and psychiatric disorders and to the clinical response to psychotherapeutics. The present study was undertaken to determine whether chronic administration of tranylcypromine (TCP), an antidepressant, and chronic stress influence GABA(B) receptor function in rat brain. The results indicate that TCP treatment, but not stress, increases GABA(B) receptor activity in the cerebral cortex, as measured by baclofen-stimulated GTPgammaS binding. In addition, chronic administration of TCP enhances significantly the locomotor response to a single dose of amphetamine, an effect that is abolished by restraint stress. These results indicate that although TCP administration modifies brain GABA(B) receptor activity, which may contribute to the antidepressant response to this agent, this effect is unrelated to the interaction of stress and TCP treatment on the locomotor response to amphetamine.

Long-term quality of life and neuropsychologic functioning for patients with CNS germ-cell tumors: from the First International CNS Germ-Cell Tumor Study.

This study evaluated the quality of life and neuropsychologic functioning among patients enrolled between 1989 and 1993 in the First International CNS Germ-Cell Tumor Study. Quality-of-life questionnaires (Short Form-36 or Child Health Questionnaire) were completed on 43 patients at median follow-up of 6.1 years after diagnosis (range, 4.5-8.8 years), and intellectual and academic testing was performed on 22 patients. Psychosocial and physical functioning of patients aged 19 years and older at follow-up was within the average range, whereas the same functioning for patients aged 18 years and younger, as reported by their parents at follow-up, was low average and borderline, respectively. Overall psychosocial and physical health summary scores were positively correlated with age at diagnosis for both groups combined. Those who received CNS radiation therapy (n = 29) reported significantly worse physical health, but similar psychosocial health, compared with those treated without radiation. Neuropsychologic testing indicated full-scale and verbal IQ, reading, spelling, and math skills in the average range, and performance IQ in the low average range. Intelligence and math skills were positively correlated with age at diagnosis. Those with germinomas significantly outperformed those with nongerminomatous/ mixed tumors on all neuropsychological measures administered. Younger patients diagnosed with CNS germ-cell tumors are at increased risk for psychosocial and physical problems as well as neuropsychologic deficits. Exposure to irradiation adversely affects overall physical functioning, whereas tumor pathology appears to be a salient neurocognitive risk factor. Collaborative and randomized studies are required to further elucidate the late effects arising from factors such as age at diagnosis, tumor histology, level of irradiation therapy, and chemotherapy toxicity among these young and potentially curable patients.

Glial somatostatin-14 expression in the rat pituitary intermediate lobe: a possible neurotrophic function during development?

Somatostatin-14 was first detected on gestational day 17 in radially-oriented, bipolar cells spanning the width of the intermediate lobe of the rat pituitary. Cells were prominent, and constituted approximately 50% of the lobe area. The presence of vimentin, the cellular shape, and the localization identified these cells as glia. At postnatal day 6, somatostatin-14 and vimentin staining appeared in stellate-shaped cells. This is in agreement with the change from bipolar to stellate shape these glia undergo after the onset of innervation ([13] Gary et al. Int. J. Devl. Neurosci. 13, 555-565, 1995). Glia were more abundant, relative to melanotropes, throughout embryonic and early postnatal development compared to adulthood. Reverse transcription-polymerase chain reaction data showed a high level of prosomatostatin mRNA in the intermediate lobe, compared to the anterior and neural lobes from postnatal day 2 animals, and a significant drop in intermediate lobe content in the adult. The correlation between the number of glia and high expression of somatostatin in neonatal relative to adult tissue, together with the close apposition of incoming axons to the abundant, radially oriented glia during innervation of the lobe, support a neurotrophic function of glia-derived somatostatin.

Effects of acute restraint stress on tyrosine hydroxylase mRNA expression in locus coeruleus of Wistar and Wistar-Kyoto rats.

Norepinephrine (NE) is thought to play a role in the stress response, and may be involved in stress-related psychopathological conditions such as depression or anxiety. Heterogeneity in individual responses to the same stressor suggest that a genetic susceptibility to the effects of stress may contribute to such pathology. To address possible mechanisms underlying this genetic aspect of the stress response, we examined acute stress-induced changes in mRNA expression for several components of the NE system in the locus coeruleus (LC) and adrenal medullae of stress-susceptible Wistar-Kyoto (WKY) rats and their parent Wistar (W) strain. Expression of tyrosine hydroxylase (TH), NE transporter (NET) and alpha(2A) receptor mRNA were measured in the LC by in situ hybridization 30 min and 2 h after the onset of 30 min restraint stress. Adrenal TH mRNA was measured by slot blots. No basal differences were observed for any measure, but in the LC, expression of TH mRNA increased by 40% in W rats at 30 min (n=8, p<0.05) and returned toward baseline by 2 h, while WKY rats showed only a non-significant 29% increase at 2 h. In contrast, adrenal TH mRNA expression increased in WKY rats at 2 h (n=3, p<0.05), with no significant change in W rats. NET and alpha(2A) mRNA were unaltered by restraint stress in both strains. Differences in the stress-reactivity of TH gene expression in the central and peripheral noradrenergic systems may be related to differences in behavioral coping strategies and autonomic responsivity to stress in these strains, and suggest that differences in noradrenergic reactivity may contribute to genetic susceptibility to stress-related pathology.

Changes in tyrosine hydroxylase mRNA expression in the rat locus coeruleus following acute or chronic treatment with valproic acid.

Valproate has proven effective in treating bipolar disorder. Though some biochemical effects of valproate are rapid, mood-stabilizing effects can take weeks, suggesting that regulatory changes in gene expression in brain neurotransmitter systems may be involved. Given a presumed role for norepinephrine (NE) in bipolar disorder, as well as the actions of mood-stabilizing drugs, we examined changes in mRNA expression for tyrosine hydroxylase (TH), the NE transporter (NET) and alpha 2A autoreceptor in the rat locus coeruleus after valproate treatment. TH mRNA increased slightly (16%) following acute treatment, and more so after chronic valproate treatment (26%), while neither NET nor alpha 2A mRNA expression changed. Further, chronic valproate treatment attenuated the elevation in TH mRNA expression induced in the LC in response to acute restraint stress. Both acute and chronic valproate treatment attenuated restraint stress-induced elevations in plasma ACTH secretion. These observations suggest that the therapeutic effects of valproate may involve regulatory alterations in TH message expression in the brain, and attenuation of stress-reactivity of the central noradrenergic system and the hypothalamic-pituitary-adrenal axis.

Have we forgotten the base rate problem? Methodological issues in the detection of distortion.

The search for valid and reliable methods of detecting malingering and distortion has become an increasingly important task for forensic psychologists and neuropsychologists. This report highlights several important methodological issues commonly observed in research on the prediction of malingering. The choice of indices for determining optimal cutoff scores on the utility of existing measures, the impact of base rates of malingering on the accuracy of prediction models, the incremental accuracy of combining multiple measures, and the relationship of test validity to the interpretation of data are described with regard to the prediction of malingering on neuropsychological tests. These methodological concerns are discussed in reference to several recent publications assessing the utility of various methods for the detection of malingering.

Expression of alpha1D adrenergic receptor messenger RNA in oxytocin- and corticotropin-releasing hormone-synthesizing neurons in the rat paraventricular nucleus.

The paraventricular nucleus of the hypothalamus contains a number of intermingled populations of neuroendocrine cell groups involved in the hormonal stress response, including cells synthesizing corticotropin-releasing hormone and oxytocin. Ascending noradrenergic afferents to the paraventricular nucleus, acting through alpha1 adrenergic receptors, are thought to play a role in stress-induced activation of the hypothalamic-pituitary-adrenal axis. We have previously demonstrated that, of the three known alpha1 adrenergic receptor subtypes, messenger RNA for the alpha1D subtype is the most prominently expressed in the paraventricular nucleus. Thus, regulation of the expression of this receptor may be important in modulation of the stress response. It is currently unknown, however, which populations of stress-related neuroendocrine cells in the paraventricular nucleus express alpha1 receptors, or whether the excitatory influence of norepinephrine in stress is exerted directly on neurons expressing oxytocin or corticotropin-releasing hormone. Thus, in the present study, we used dual in situ hybridization, combining a digoxigenin-labeled riboprobe encoding the rat alpha1D adrenergic receptor with radiolabeled riboprobes for oxytocin or corticotropin-releasing hormone, to determine the degree to which these neurons in the paraventricular nucleus express alpha1D adrenergic receptors. In sections through the rostral and mid-level paraventricular nucleus, nearly all (>95%) oxytocin neurons also expressed alpha1D messenger RNA. In contrast, the populations of corticotropin-releasing hormone- and alpha1D-expressing cells overlapped only partially, with most alpha1D expression situated more laterally. A subset (37%) of the neurons expressing corticotropin-releasing hormone also expressed alpha1D messenger RNA, and these were found almost entirely within the region of overlap in the lateral aspect of the medial parvocellular region. These observations support a direct role for alpha1 receptors in regulation of oxytocin secretion. Expression of alpha1D messenger RNA in distinct subsets of cells synthesizing corticotropin-releasing hormone may also help to clarify contradictory and inconsistent observations in the literature regarding the role of norepinephrine in the stress response, and may account for a presumed stressor-specific role for norepinephrine in activation of the hypothalamic-pituitary-adrenal axis.

Pilot neuropsychological findings from a treatment regimen consisting of intensive chemotherapy and bone marrow rescue for young children with newly diagnosed malignant brain tumors.

Ten children (6 girls and 4 boys) who completed a protocol in which their localized brain tumors were successfully treated without cranial irradiation were referred for neuropsychological assessment. At the time of testing, they were disease free without any neuroaxis dissemination or leptomeningeal disease. Tumor types included pineoblastoma, glioblastoma, ependymoma, PNET and medulloblastoma. They had a mean age of 5 years and 8 months (SD = 1.86; range = 2.1-8.9 years) and were an average of 37.8 months post bone marrow transplant (SD = 16.42; range = 14-58 months). Neuropsychological data from this study reveal that the mean scores for this nonradiated group of children were within the average range for the following domains: academic achievement tests of reading, spelling and mathematics, verbal and visual memory, visual-motor integration, social-emotional and behavioral functioning. Furthermore, this group of children were performing within the low average range of overall Intelligence, as well as both verbal IQ/verbal reasoning and performance IQ/abstract visual reasoning. On tasks of fine motor dexterity, this group was within the low average range when using their dominant hand; however, they performed within the borderline range when using their non-dominant hand. Of note, this group of children demonstrated significant deficits within the borderline to impaired ranges on language tasks of expressive picture naming and receptive picture vocabulary.

Differential innervation of individual melanotropes suggests a role for nonsynaptic inhibitory regulation of the developing and adult rat pituitary intermediate lobe.

Dopamine and GABA were detected in intermediate lobe axons around birth, and early axons were closely apposed to glial cells and processes, possibly using them for guidance. In the adult, axons containing colocalized dopamine and GABA were distributed in a distinct pattern within the lobe, with plexuses located dorsally and ventrally. Axons preferentially followed glial processes in interlobular septa, yet were also interspersed between melanotropes. Individual melanotropes were contacted by varying numbers of axon terminals, with some devoid of contacts. Boutons contained both small clear vesicles and large dense-cored vesicles; membrane specializations were not well-developed. From these findings we concluded that in addition to direct synaptic inhibition, dopamine and GABA could stimulate their receptors by mechanisms similar to "parasynaptic" [Schmitt (1984) Neuroscience, 13:991-1001] or "volume" [Agnati et al. (1995) Neuroscience, 69:711-726] transmission as proposed for the CNS. Humoral agents passing into the intermediate lobe from portal vessels, thus acting as classical hormones, further regulate the melanotropes. Moreover, approximately 50% of the axonal elements were closely apposed to glia, suggesting that glia could have regulatory roles. Previous studies from our laboratory [Chronwall et al. (1987) Endocrinology, 120:1201-1211; Chronwall et al. (1988) Endocrinology, 123:1992:1202] demonstrated heterogeneity in proopiomelanocortin (POMC) biosynthesis among individual melanotropes, prompting the hypothesis that the degree of innervation could govern the expression of certain molecules. We combined immunohistochemistry and in situ hybridization histochemistry to evaluate whether melanotrope molecular heterogenity is spatially correlated with axons and terminals. Tentatively, melanotropes expressing low levels of POMC and alpha1A subunit P/Q type Ca2+ channel mRNAs often were apposed to axons, whereas those with low levels of D2L receptor mRNA rarely were contacted by axons, suggesting that innervation could be one of the factors inducing and maintaining heterogeneity.

Dopamine D2 receptor effects on GABA(A) receptor expression may modify melanotrope peptide release.

Stimulation of melanotrope dopamine D2 receptors decreases mitotic rate, calcium channel activity, and the biosynthesis of several proteins. This study demonstrates that D2 receptor activation also affects GABA(A) receptor beta2/beta3 subunit immunoreactivity. Following chronic treatment with haloperidol, a D2 receptor antagonist, GABA(A) receptor immunoreactivity increased, whereas it decreased after chronic treatment with bromocriptine, a dopamine D2 receptor agonist. Thus, these data indicate that D2 function regulates GABA(A) receptor expression in melanotropes, a mechanism by which peptide release may be modified.

G-protein expression in melanotropes changes coincident with innervation of the developing rat pituitary intermediate lobe.

The two isoforms of the dopamine D2 receptor, the D2short and the D2long differ in a 29 amino acid insert in the third cytoplasmic loop with which G proteins interact. We have previously reported that in rat melanotropes, expression of D2short increases markedly at the end of the first postnatal week which is concurrent with innervation of the intermediate lobe. Using immunohistochemistry, this study examined expression of G alpha i1/2, G alpha i3, G alpha o and G alpha s proteins before and after dopaminergic innervation. G alpha i3 increased through gestational day 20, and then remained level to postnatal day 6. At this time, coinciding with the induction of D2short expression, G alpha i3 immunoreactive intensity increased markedly, possibly indicating co-regulation of these proteins. On postnatal day 6, G alpha s immunoreactive intensity increased in some, but not all, melanotropes. The resulting heterogeneity in Gs expression persisted in the adult. G alpha i1/2 immunoreactivity did not change and G alpha o was detected only subsequent to the event of innervation. Thus, dopamine released from axons and acting through D2 receptor stimulation could increase G alpha i3 immunoreactivity and decrease G alpha s immunoreactive intensity in some melanotropes.

Dopamine D2 receptor stimulation alters G-protein expression in rat pituitary intermediate lobe melanotropes.

Stimulation of dopamine D2 receptors inhibits melanotrope pro-opiomelanocortin (POMC) biosynthesis and alpha-melanocyte-stimulating hormone (MSH) secretion. These effects are mediated by G-protein alpha i- and alpha o-subunits and are reversed by stimulating receptors linked to activation of G alpha s protein. Melanotrope activity is increased by haloperidol, a D2 receptor antagonist, and decreased by bromocriptine, a D2 receptor agonist. Both the short and long isoforms of the D2 receptor mRNA and protein increase following chronic haloperidol treatment. After chronic bromocriptine treatment the short isoform is downregulated, whereas the long isoform is upregulated. Our hypothesis is that specific G protein alpha- subunits alter in pattern of expression similarly to the receptor isoforms. Using immunohistochemistry and in situ hybridization, this study examined changes in G alpha i, G alpha o, and G alpha s protein and mRNA expression following chronic treatments with bromocriptine or haloperidol. G alpha i3 and G alpha o immunoreactivities increased following bromocriptine treatment, whereas G alpha s and G alpha i1/2 did not change. Gs immunoreactivity increased after haloperidol treatment, whereas G alpha i1/2, G alpha i3, and G alpha o did not change. G alpha i and G alpha o mRNA increased following bromocriptine and decreased following haloperidol treatments, whereas the inverse results were observed with G alpha s mRNA. These results suggest D2 receptor activation can specifically increase G alpha i3 and G alpha o expression, and D2 receptor blockade increases G alpha s expression.

Lactation and salt loading similarly alter neuropeptide Y, but differentially alter somatostatin, in separate sets of rat neural lobe axons.

Neuropeptide Y (NPY) and somatostatin immunoreactivities are present in neural lobe axons of the rat pituitary. Both peptides are upregulated during lactation, because NPY gene expression increases in the hypothalamus and plasma concentrations of somatostatin are elevated. However, the effects of lactation on NPY and somatostatin in the neural lobe are unknown. Although NPY immunoreactivity increases in the neural lobe following salt loading of male rats, the somatostatin response is unknown. To answer these questions, NPY and somatostatin immunoreactivities in the neural lobe were examined during lactation and salt loading using immunohistochemistry and image analysis. On day 2 of lactation, the area covered by immunoreactivity, a combined measurement of axon density and size of axonal swellings, of both NPY and somatostatin increased compared to ovariectomized rats. The increase in NPY was four- to fivefold greater than that of somatostatin. By day 10 of lactation, values returned to those of ovariectomized rats. Following 10 days of salt loading, the area covered by NPY immunoreactivity increased approximately 10-fold over control male rats, whereas somatostatin remained unchanged. NPY and somatostatin were not colocalized in neural lobe axons in either paradigm, demonstrating that two different neuronal populations were involved in both cases. These data indicate that NPY and somatostatin were regulated similarly during lactation, but differentially following salt loading.

Types and activities of voltage-operated calcium channels change during development of rat pituitary neurointermediate lobe.

Cultures of pituitary neurointermediate lobe cells were established from rats aged 1, 12, and 42 days to identify the types and assess the activities of Ca2+ channels present in melanotropes, glial-like cells, and fibroblasts during development. Day 12 represents the time at which dopaminergic axons have become distributed throughout the lobe, glial cells begin to lose their radial orientation, and melanotropes robustly express the short isoform of the dopamine D2 receptor. Thus, we studied Ca2+ channels in relation to the event of innervation of melanotropes. Real-time fluorescence video microscopy, in the presence of pharmacological agents, which block L-, N-, P-, and T-type channels, was used as an indirect measurement of channel activity. Assessment of cell type was verified by triple-label fluorescence immunohistochemistry. In melanotropes, extracellular Ca2+ addition caused Ca2+ influx through omega-conotoxin GVIA-sensitive, N-type channels on days 1 and 12 but not on day 42. The K+ depolarization induced an increase in intracellular Ca2+ concentration in all age-groups. This effect was decreased by nifedipine, an L-type channel blocker, at all ages, and by omega-agatoxin IVa, a P-type blocker, only on day 42. These results demonstrate that the predominance of N- or P-type channels on melanotropes is age-dependent and can be correlated with other developmental changes. The T-type blocker, NiSO4, had no effect. In glial-like cells of all ages, extracellular Ca2+ addition resulted in an increase in intracellular Ca2+ concentration, which was inhibited only by NiSO4. The percentage of responsive glial-like cells was equally high in days 1 and 12 cultures, then declined by day 42. The K+ depolarization had no effect on glial-like cells. Fibroblasts did not respond significantly to extracellular Ca2+ or K+ depolarization, indicating little detectable activity by this methodology from functional voltage-operated Ca2+ channels.

Melanotrope dopamine D2 receptor isoform expression in the developing rat pituitary.

This study measured melanotrope mRNA and protein expression for the dopamine D2 receptor, and its long isoform, in relation to the appearance of dopamine in axons of the postnatal rat pituitary intermediate lobe. At postnatal day 2, prior to the onset of dopaminergic innervation, D2 receptor (D2T) mRNA was expressed heterogeneously in a subpopulation of melanotropes which also expressed the long isoform (DL). The D2L mRNA appeared to be predominant during early postnatal development, since the D2T probe, which did not discriminate between the isoforms, and the D2L probe hybridized generally to the same cells, as demonstrated in serial sections. Immunohistochemical methods, using two different antisera for the D2T receptor, however, indicated a low level of protein in most melanotropes. Localization of D2L protein corresponded well to D2T receptor mRNA distribution. At day 10, representing a time when dopamine is present in axons throughout the lobe, both D2T receptor mRNA and protein were detected in a significantly larger population of melanotropes than those expressing D2L mRNA and protein. This suggests the appearance of detectable short isoform (D2S) mRNA in virtually all melanotropes and implicates dopamine as a possible signal for increasing D2S isoform mRNA expression.