The Small Interactor of PKD2 protein promotes the assembly and ciliary entry of the Chlamydomonas PKD2-mastigoneme complexes.

In Chlamydomonas, the channel polycystin 2 (PKD2) is primarily present in the distal region of cilia, where it is attached to the axoneme and mastigonemes, extracellular polymers of MST1. In a smaller proximal ciliary region that lacks mastigonemes, PKD2 is more mobile. We show that the PKD2 regions are established early during ciliogenesis and increase proportionally in length as cilia elongate. In chimeric zygotes, tagged PKD2 rapidly entered the proximal region of PKD2-deficient cilia, whereas the assembly of the distal region was hindered, suggesting that axonemal binding of PKD2 requires de novo assembly of cilia. We identified the protein Small Interactor of PKD2 (SIP), a PKD2-related, single-pass transmembrane protein, as part of the PKD2-mastigoneme complex. In sip mutants, stability and proteolytic processing of PKD2 in the cell body were reduced and PKD2-mastigoneme complexes were absent from the cilia. Like the pkd2 and mst1 mutants, sip mutant cells swam with reduced velocity. Cilia of the pkd2 mutant beat with an increased frequency but were less efficient in moving the cells, suggesting a structural role for the PKD2-SIP-mastigoneme complex in increasing the effective surface of Chlamydomonas cilia.

Engaging homeostatic plasticity to treat depression.

Major depressive disorder (MDD) is a complex and heterogeneous mood disorder, making it difficult to develop a generalized, pharmacological therapy that is effective for all who suffer from MDD. Through the fortuitous discovery of N-methyl-D-aspartate receptor (NMDAR) antagonists as effective antidepressants, we have gained key insights into how antidepressant effects can be produced at the circuit and molecular levels. NMDAR antagonists act as rapid-acting antidepressants such that relief from depressive symptoms occurs within hours of a single injection. The mode of action of NMDAR antagonists seemingly relies on their ability to activate protein-synthesis-dependent homeostatic mechanisms that restore top-down excitatory connections. Recent evidence suggests that NMDAR antagonists relieve depressive symptoms by forming new synapses resulting in increased excitatory drive. This event requires the mammalian target of rapamycin complex 1 (mTORC1), a signaling pathway that regulates synaptic protein synthesis. Herein, we review critical studies that shed light on the action of NMDAR antagonists as rapid-acting antidepressants and how they engage a neuron's or neural network's homeostatic mechanisms to self-correct. Recent studies notably demonstrate that a shift in γ-amino-butyric acid receptor B (GABABR) function, from inhibitory to excitatory, is required for mTORC1-dependent translation with NMDAR antagonists. Finally, we discuss how GABABR activation of mTORC1 helps resolve key discrepancies between rapid-acting antidepressants and local homeostatic mechanisms.

Rapid antidepressants stimulate the decoupling of GABA(B) receptors from GIRK/Kir3 channels through increased protein stability of 14-3-3η.

A single injection of N-methyl-D-aspartate receptor (NMDAR) antagonists produces a rapid antidepressant response. Lasting changes in the synapse structure and composition underlie the effectiveness of these drugs. We recently discovered that rapid antidepressants cause a shift in the γ-aminobutyric acid receptor (GABABR) signaling pathway, such that GABABR activation shifts from opening inwardly rectifiying potassium channels (Kir/GIRK) to increasing resting dendritic calcium signal and mammalian Target of Rapamycin activity. However, little is known about the molecular and biochemical mechanisms that initiate this shift. Herein, we show that GABABR signaling to Kir3 (GIRK) channels decreases with NMDAR blockade. Blocking NMDAR signaling stabilizes the adaptor protein 14-3-3η, which decouples GABABR signaling from Kir3 and is required for the rapid antidepressant efficacy. Consistent with these results, we find that key proteins involved in GABABR signaling bidirectionally change in a depression model and with rapid antidepressants. In socially defeated rodents, a model for depression, GABABR and 14-3-3η levels decrease in the hippocampus. The NMDAR antagonists AP5 and Ro-25-6981, acting as rapid antidepressants, increase GABABR and 14-3-3η expression and decrease Kir3.2. Taken together, these data suggest that the shift in GABABR function requires a loss of GABABR-Kir3 channel activity mediated by 14-3-3η. Our findings support a central role for 14-3-3η in the efficacy of rapid antidepressants and define a critical molecular mechanism for activity-dependent alterations in GABABR signaling.

Small Interactor of PKD2 (SIP), a novel PKD2-related single-pass transmembrane protein, is required for proteolytic processing and ciliary import of Chlamydomonas PKD2.

In Chlamydomonas cilia, the ciliopathy-relevant TRP channel PKD2 is spatially compartmentalized into a distal region, in which PKD2 binds the axoneme and extracellular mastigonemes, and a smaller proximal region, in which PKD2 is more mobile and lacks mastigonemes. Here, we show that the two PKD2 regions are established early during cilia regeneration and increase in length as cilia elongate. In abnormally long cilia, only the distal region elongated whereas both regions adjusted in length during cilia shortening. In dikaryon rescue experiments, tagged PKD2 rapidly entered the proximal region of PKD2-deficient cilia whereas assembly of the distal region was hindered, suggesting that axonemal docking of PKD2 requires de novo ciliary assembly. We identified Small Interactor of PKD2 (SIP), a small PKD2-related protein, as a novel component of the PKD2-mastigoneme complex. In sip mutants, stability and proteolytic processing of PKD2 in the cell body were reduced and PKD2-mastigoneme complexes were absent from mutant cilia. Like the pkd2 and mst1 mutants, sip swims with reduced velocity. Cilia of the pkd2 mutant beat with normal frequency and bending pattern but were less efficient in moving cells supporting a passive role of the PKD2-SIP-mastigoneme complexes in increasing the effective surface of Chlamydomonas cilia.

An Innovative Approach to Care: Integrating Mental Health Services Through Telemedicine in Rural School-Based Health Centers.

In rural communities, primary care providers continue to provide mental health services, and about 70% of children and adolescents identified to have a psychiatric disorder never receive treatment. A telehealth model for providing integrated mental health services in a school-based health clinic has the potential to increase access to specialized care for the most vulnerable youths. This column provides an overview of the strategies used to implement and integrate such a model in West Virginia. Operationalization, barriers, challenges, and judicious resource use are discussed. Appropriate reimbursement for services and state-specific legislation to ensure consistent revenue to sustain the program are considered.

The effect of serum and experimental variables on the transferrin and reticulocyte interaction.

The transfer of iron from transferrin to the developing erythrocyte is a research area of high interest and considerable controversy. We have found that the results of transferrin-reticulocyte incubation studies are quite sensitive to the experimental procedures that are utilized. Reticulocytosis has been induced in rabbits by phelbotomy and phenylhydrazine injections. While the latter gives a higher reticulocyte count, the cells appear to exhibit an altered transferrin-membrane interaction. Transferrin has been iodinated by published methods utilizing chloramine-T and molecular iodine. The iodotransferrin products exhibit the same iron donation ability, however, evidence was found that the chloramine-T treatment leads to a nonspecific binding of transferrin to the reticulocyte. The means of saturating transferrin with 59Fe is also of prime importance. Fe(NH4)2(SO4)2 and especially FeCl3 were found to yield nonspecifically bound iron when added to transferrin or serum. This artifact was reflected in an altered transferrin-reticulocyte interaction. Using what we believe to be optimal conditions, the effect of serum on the transferrin-reticulocyte system was re-examined. The results clearly indicated an enhancement of iron uptake by reticulocytes in the presence of serum, as well as an accelerated incorporation of iron by the cytoplasmic fraction.

On the preparation of bovine alpha-thrombin.

An improved method for the preparation of bovine alpha-thrombin is described. The procedure involves that activation of partially purified prothrombin with tissue thromboplastin followed by chromatography on Sulfopropyl-Sephadex C-50. The purified enzyme is homogeneous on polyacrylamide discontinuous gel electrophoresis and has a specific activity toward fibrinogen of 2.200-2,700 N.I.H. U/mg. Its stability on storage in liquid media is dependent on both ionic strength and temperature. Increasing ionic strength and decreasing temperature result in optimal stability. The denaturation of alpha-thrombin by guanidine hydrochloride was found to be a partially reversible process with the renatured species possessing properties similar to "aged" thrombin. In addition, the catalytic properties of alpha-thrombin covalently attached to agarose gel beads were also examined. The activity of the immobilized enzyme toward fibrinogen was affected to a much greater extent than was the hydrolysis of low molecular weight, synthetic substrates.

Atypical antidepressants versus imipramine in the treatment of major depression: a meta-analysis.

BACKGROUND: Our investigation involved a quantitative literature review technique known as meta-analysis to compare the efficacy of three newer antidepressants and imipramine. METHOD: We examined seven major journals in psychiatry from 1980 through 1990, inclusive, and selected those investigations of imipramine, trazodone, bupropion, and fluoxetine that met our minimal criteria for interpretability. These criteria included: (1) the presence of a placebo control, (2) double-blind status, (3) the use of the Hamilton Rating Scale for Depression as a dependent variable measure, (4) the use of nongeriatric adults with a diagnosis of major depression by DSM or RDC standards, and (5) the presence of reported means and standard deviations in the investigation, or sufficient data that allowed such to be computed. Each study of four antidepressants was analyzed for an effect size of the drug investigated. The effect size allows for a determination of the efficacy of a particular drug as compared with placebo, measured in standard deviation units. RESULTS: The data indicated that all four agents are effective as compared with placebo. Furthermore, there is no evidence that the newer heterocyclic agents are less effective than imipramine, as an ANOVA showed no statistically significant difference between the effect sizes of the four antidepressants. CONCLUSION: These data are discussed in terms of characteristics of the various investigations and the need for further research comparing the efficacy of psychopharmacologic agents.

Affinity chromatography of platelets on immobilized thrombin: retention of catalytic activity by platelet-bound thrombin.

Radioactivity from I125-labeled human platelets was measured to estimate the extent of binding of platelet surface proteins to immobilized thrombin. 1-3% of the radioactivity was bound with 10-20% of this amount apparently irreversibly bound to the thrombin matrix. Site-specific chemical modification of thrombin with pyridoxal-5'-phosphate, N-bromosuccinimide or tetranitromethane resulted in a variable reduction of the amount of radiolabel bound. When thrombin modified with H-D-PheProArg-chloromethyl ketone (PPACK) was coupled to the matrix, there was no difference in the binding of platelet membrane proteins when compared to a control thrombin preparation while thrombin modified with tosyl-Lys-chloromethyl ketone (TLCK) coupled to the matrix did not bind radiolabel any more effectively than albumin which served as the control. However, when thrombin was modified with PPACK after coupling to the agarose matrix, ability to bind radiolabel was lost. Thrombin bound to platelets remained catalytically active when assayed with a peptide nitroanilide substrate. These results suggest tight binding between thrombin and platelets that is not only not dependent on active site integrity but leaves the bound thrombin catalytically competent.

Mobilization of iron from specifically labeled reticulocyte ghosts.

Specifically labeled 59Fe ghosts have been prepared by incubation of whole reticulocytes with 59Fe3+-transferrin-CO3(2)-- followed by washing and ghost isolation. The binding of 59Fe by the membrane fraction is quite stable over a wide range of conditions, but iron mobilization occurs on incubation with chelating agents or cell lysate. The time course of 59Fe mobilization by unlabeled reticulocyte lysate exhibits five apparently zero-order phases. The rate of iron mobilization is linearly dependent on the concentration of 59Fe ghosts present in the incubation mixture. In contrast, the relative concentration of lysate appears to exhibit a saturation dependence with regard to membrane iron mobilization. Bathophenanthroline sulfonate follows a multiphasic time course of iron mobilization similar to that found with the lysate. Lysate from mature erythrocytes was found to mobilize iron with kinetics that are identical to reticulocyte lysate. The number and duration of the phases is independent of the mobilizing agent. The role of the membrane fraction in regulating the rate of iron release to cytosol was also investigated by the repetitive incubation of 59Fe ghosts with fresh lysate. The rate of 59Fe mobilization depended on the condition of the ghost with regard to prior 59Fe depletion. This publication emphasizes the active role of the membrane fraction in determining the rate at which iron will become available to the cytosol and the possibility that cytosol factors modulate the action of membrane bound components.

A 30-year progress report on a VA satellite psychiatric clinic program.

This paper describes a program of rural satellite psychiatric clinics that has evolved within a Veterans Affairs medical center over the past three decades. The eight clinics are staffed by a single team that includes a psychiatrist, a psychiatric nurse, and two mental health technicians. The team travels to the eight sites at least once each month. The program has achieved its goal of providing outpatient screening, diagnosis, and treatment services to veterans who would otherwise have no access to specialty psychiatric care due to distance and transportation problems.

Analyze: a comprehensive, operator-interactive software package for multidimensional medical image display and analysis.

A comprehensive software package, called ANALYZE, has been developed (1) which permits detailed investigation and evaluation of multidimensional biomedical images. ANALYZE can be used with 3-D imaging modalities based on x-ray computed tomography, radionuclide emission tomography, ultrasound tomography, and magnetic resonance imaging. The software is written entirely in "C" and runs on standard UNIX workstations. The ANALYZE package features integrated, complimentary tools for fully interactive display, manipulation and measurement of multidimensional image data. The software architecture permits systematic enhancements and upgrades which has fostered development of a readily expandable package. It provides an effective shell for custom software prototyping and turnkey applications. This paper provides a general description of this software as well as specific details on the methodology employed to develop it, both conceptual and technical. Applications of the software are illustrated.

Guiding parents through the death of their infant.

Supporting a family through the impending death of their infant challenges the health care team to create opportunities for the parents to complete the attachment process and begin to grieve. Even in the neonatal intensive-care unit, the family can be brought together with their child in activities that comfort and console, lay the foundation for positive memories, and initiate healing and closure after the death of the infant. Interventions for grieving parents are described in this article.

Psychoneuroimmunology: where are we, where are we going?

We have reviewed briefly the current status of research on central nervous system-immune system interactions, focusing attention on the neural and humoral pathways by which CNS and IS communicate and interact and on the effects of stress and psychiatric illness on immune function. It is evident that CNS-IS communication occurs by direct innervation of lymphoid organs and by means of hormones, neuropeptides and cytokines. There is also clear evidence that humoral substances each of which were thought to be the product of one specific cell type are elaborated and secreted by a variety of cell types. This observation suggests a new unified concept of CNS-IS interactions with mediators of these interactions being produced ubiquitously and acting on cells of the two systems. In examining the effects of stress on IS it has become apparent that stress of various types can have a depressive effect on immune functions, primarily at the level of T lymphocytes and NK cells. This suggests that the defense mechanisms affected by stress are those which are responsible for cytotoxic effector responses. These findings are interesting in that they support older studies implicating stress in the pathogenesis and/or the clinical course of neoplastic diseases. Further support for a role of stress-induced immunodepression in morbidity comes from a very interesting, recent prospective study showing that stress will affect susceptibility to viruses. Finally, exploration of the mechanisms of stress-induced immunodepression, suggests that a variety of mediators which regulate lymphocyte interactions and activation may be affected, perhaps at the level of gene expression.(ABSTRACT TRUNCATED AT 250 WORDS)

mTORC1-dependent protein synthesis underlying rapid antidepressant effect requires GABABR signaling.

Administration of N-methyl-D-aspartate receptors (NMDAR) antagonists initiates a rapid anti-depressant response requiring mammalian Target of Rapamycin Complex 1 (mTORC1) kinase; however the molecular mechanism is unknown. We have determined that upon NMDAR blockade, dendritic γ-amino-butyric acid B receptors (GABABR) facilitate dendritic calcium entry. The GABABR-mediated increase in calcium signal requires the availability of dendritic L-type calcium channels. Moreover, GABABR can activate mTOR and increase mTOR dependent expression of BDNF under the same NMDAR blocked conditions. In vivo, blocking GABABR prevents the fast-acting, anti-depressant effect of the NR2B antagonist, Ro-25-6891, decreases active mTORC1 kinase, and reduces expression of BDNF and the AMPA receptor subunit GluA1. These findings propose a novel role for GABABRs in the antidepressant action of NR2B antagonists and as an initiator/regulator of mTORC1-mediated translation.