The effectiveness and safety of dialectical behavior therapy for suicidal ideation and behavior in autistic adults: a pragmatic randomized controlled trial.

BACKGROUNDS: Many autistic people in mental health are suicidal. This study evaluated the effectiveness of dialectical behavior therapy (DBT) v. treatment as usual (TAU) in reducing suicidal ideation and suicide attempts. METHODS: At six Dutch mental health centers, 123 outpatients (18-65 years) with DSM-5 diagnosed autism spectrum disorder (ASD) and suicidal behavior were randomly assigned to the DBT intervention group (n = 63) or TAU control group (n = 60). Assessments were conducted at baseline, post-treatment at 6 months and 12-month follow-up. The primary outcomes were severity of suicidal ideation and frequency of suicide attempts. The severity of depression and social anxiety were secondary outcomes. RESULTS: At end-of-treatment, DBT significantly reduced both suicidal ideation (z = -2.24; p = 0.025; b = -4.41; s.e. = 197.0) and suicide attempts (z = -3.15; p = 0.002; IRR = 0.046; s.e. = 0.045) compared to TAU, but lost statistical significance at the 12-month follow-up. Depression severity significantly decreased with DBT (z = -1.99; p = 0.046: b = -2.74; s.e. = 1.37) remaining so at 12 months (z = -2.46; p = 0.014; b = -3.37; s.e. = 1.37). No effects were observed on social anxiety. Severe adverse events included two suicides in the TAU condition. CONCLUSIONS: DBT is an acceptable, safe, and short-term effective intervention to reduce suicidal ideation and suicide attempts in autistic adults with suicidal behavior.

[Treatment of emotion regulation problems in people with neurofibromatosis type 1]. / Behandeling van emotieregulatieproblemen bij mensen met neurofibromatose type 1.

Individuals with the genetic disorder neurofibromatosis type 1 (NF1) are typically diagnosed in a medical hospital setting strongly relying on the presence of well-defined physical symptoms such as neurofibromas or pigmentary spots (known as café-au-lait spots). In mental health care settings, however, aside from a few highly specialized centres, the diagnosis and treatment of individuals with NF1 receives little attention, while the need for psychological treatment is increasingly identified, both in clinical practice and in the scientific literature. Occasional referrals of individuals with NF1 to the mental health services are often only targeted at psychological assessment. Subsequent treatment, however, is usually lacking. We describe two individuals with NF1 for whom by means of specialized clinical neuropsychological assessment, participation in a tailored dialectical behavior therapy (DBT) skills training was indicated. We exposit how they were able to develop their skills and how they themselves and their significant others experienced the treatment.

Necrosome-positive granulovacuolar degeneration is associated with TDP-43 pathological lesions in the hippocampus of ALS/FTLD cases.

AIM: Granulovacuolar degeneration (GVD) in Alzheimer's disease (AD) involves the necrosome, which is a protein complex consisting of phosphorylated receptor-interacting protein kinase 1 (pRIPK1), pRIPK3 and phosphorylated mixed lineage kinase domain-like protein (pMLKL). Necrosome-positive GVD was associated with neuron loss in AD. GVD was recently linked to the C9ORF72 mutation in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with transactive response DNA-binding protein (TDP-43) pathology (FTLD-TDP). Therefore, we investigated whether GVD in cases of the ALS-FTLD-TDP spectrum (ALS/FTLD) shows a similar involvement of the necrosome as in AD, and whether it correlates with diagnosis, presence of protein aggregates and cell death in ALS/FTLD. METHODS: We analysed the presence and distribution of the necrosome in post-mortem brain and spinal cord of ALS and FTLD-TDP patients (n = 30) with and without the C9ORF72 mutation, and controls (n = 22). We investigated the association of the necrosome with diagnosis, the presence of pathological protein aggregates and neuronal loss. RESULTS: Necrosome-positive GVD was primarily observed in hippocampal regions of ALS/FTLD cases and was associated with hippocampal TDP-43 inclusions as the main predictor of the pMLKL-GVD stage, as well as with the Braak stage of neurofibrillary tangle pathology. The central cortex and spinal cord, showing motor neuron loss in ALS, were devoid of any accumulation of pRIPK1, pRIPK3 or pMLKL. CONCLUSIONS: Our findings suggest a role for hippocampal TDP-43 pathology as a contributor to necrosome-positive GVD in ALS/FTLD. The absence of necroptosis-related proteins in motor neurons in ALS argues against a role for necroptosis in ALS-related motor neuron death.

The effect of dialectical behaviour therapy in autism spectrum patients with suicidality and/ or self-destructive behaviour (DIASS): study protocol for a multicentre randomised controlled trial.

BACKGROUND: Many persons with autism spectrum disorder (ASD) are treated in long-term specialised care. In this population, suicidal behaviour troubles patients, families, and specialists in the field because it is difficult to treat. At present, there is no documented effective therapy for suicidal behaviour in ASD (Autism Research 7:507-521, 2014; Crisis 35:301-309, 2014). Dialectical Behaviour Therapy (DBT) is an efficacious treatment programme for chronically suicidal and/or self-harm behaviour in patients with Borderline Personality Disorder (J Psychiatry 166:1365-1374, 2014; Linehan MM. Cognitive behavioural therapy of borderline personality disorder. 1993). This study will evaluate the efficacy of DBT in persons with ASD and suicidal/ self- destructive behaviour in a multicentre randomised controlled clinical trial. METHOD: One hundred twenty-eight persons with autism and suicidal and/or self-harming behaviour will be recruited from specialised mental healthcare services and randomised into two conditions: 1) the DBT condition in which the participants have weekly individual cognitive behavioural therapy sessions and a 2.5 h skills training group session twice per week during 6 months, and 2) the treatment as usual condition which consists of weekly individual therapy sessions of 30-45 min with a psychotherapist or social worker. Assessments will take place at baseline, at post-treatment (6 months), and after a follow-up period of 12 months. The mediators will also be assessed at 3 months. The primary outcome is the level of suicidal ideation and behaviour. The secondary outcomes are anxiety and social performance, depression, core symptoms of ASD, quality of life, and cost-utility. Emotion regulation and therapeutic alliance are hypothesised to mediate the effects on the primary outcome. DISCUSSION: The results from this study will provide an evaluation of the efficacy of DBT treatment in persons with ASD on suicidal and self-harming behaviour. The study is conducted in routine mental health services which enhances the generalisability of the study results to clinical practice. TRIAL REGISTRATION: ISRCTN96632579. Registered 1 May 2019. Retrospectively registered.

Serum neurofilament light chain levels as a marker of upper motor neuron degeneration in patients with Amyotrophic Lateral Sclerosis.

AIMS: Amyotrophic lateral sclerosis (ALS) is the most common motor neuron degeneration disease with a diagnostic delay of about 1 year after symptoms onset. In ALS, blood neurofilament light chain (NfL) levels are elevated, but it is not entirely clear what drives this increase and what the diagnostic performance of serum NfL is in terms of predictive values and likelihood ratios. The aims of this study were to further explore the prognostic and diagnostic performances of serum NfL to discriminate between patients with ALS and ALS mimics, and to investigate the relationship between serum NfL with motor neuron degeneration. METHODS: The diagnostic performances of serum NfL were based on a cohort of 149 serum samples of patients with ALS, 19 serum samples of patients with a disease mimicking ALS and 82 serum samples of disease control patients. The serum NfL levels were correlated with the number of regions (thoracic, bulbar, upper limb and lower limb) displaying upper and/or lower motor neuron degeneration. The prognostic performances of serum NfL were investigated based on a Cox regression analysis. RESULTS: The associated predictive values and likelihood ratio to discriminate patients with ALS and ALS mimics were established. Serum NfL was associated with motor neuron degeneration driven by upper motor neuron (UMN) degeneration and was independently associated with survival in patients with ALS. CONCLUSIONS: Altogether, these findings suggest that elevated serum NfL levels in ALS are driven by UMN degeneration and the disease progression rate and are independently associated with survival at time of diagnosis.

[Dialectical behaviour theory in the Netherlands: implementation and consolidation]. / Dialectische gedragstherapie in Nederland: implementatie en consolidatie.

BACKGROUND: More and more evidence-based treatments for severe personality disorders are becoming available. Nevertheless, there are problems with the implementation of these treatments and it is proving difficult to keep the treatment programmes running. However, teams which offer dialectical behavior therapy (DBT) seem to survive. AIM: To find out which factors enable dbt teams to survive. METHOD: Twenty-five Dutch DBT teams received a questionnaire about factors that could be influencing the continuation of the DBT treatment programmes. The questionnaire consisted of 9 open questions, 2 multiple-choice questions and 26 closed questions. RESULTS: The results show that the continued existence of the treatment programmes is due largely to the commitment of both the team and its managers. They all feel embedded in the organisation as a whole, feel connected with one another and are supportive of the method. CONCLUSION: A well-functioning consultation team seems to be of crucial importance for the continued existence of the DBT programme. We believe that independent external supervision is essential to keep the dbt teams alert and aware of current trends and developments.

A deep phenotyping study in mouse and iPSC models to understand the role of oligodendroglia in optic neuropathy in Wolfram syndrome.

Wolfram syndrome (WS) is a rare childhood disease characterized by diabetes mellitus, diabetes insipidus, blindness, deafness, neurodegeneration and eventually early death, due to autosomal recessive mutations in the WFS1 (and WFS2) gene. While it is categorized as a neurodegenerative disease, it is increasingly becoming clear that other cell types besides neurons may be affected and contribute to the pathogenesis. MRI studies in patients and phenotyping studies in WS rodent models indicate white matter/myelin loss, implicating a role for oligodendroglia in WS-associated neurodegeneration. In this study, we sought to determine if oligodendroglia are affected in WS and whether their dysfunction may be the primary cause of the observed optic neuropathy and brain neurodegeneration. We demonstrate that 7.5-month-old Wfs1∆exon8 mice display signs of abnormal myelination and a reduced number of oligodendrocyte precursor cells (OPCs) as well as abnormal axonal conduction in the optic nerve. An MRI study of the brain furthermore revealed grey and white matter loss in the cerebellum, brainstem, and superior colliculus, as is seen in WS patients. To further dissect the role of oligodendroglia in WS, we performed a transcriptomics study of WS patient iPSC-derived OPCs and pre-myelinating oligodendrocytes. Transcriptional changes compared to isogenic control cells were found for genes with a role in ER function. However, a deep phenotyping study of these WS patient iPSC-derived oligodendroglia unveiled normal differentiation, mitochondria-associated endoplasmic reticulum (ER) membrane interactions and mitochondrial function, and no overt signs of ER stress. Overall, the current study indicates that oligodendroglia functions are largely preserved in the WS mouse and patient iPSC-derived models used in this study. These findings do not support a major defect in oligodendroglia function as the primary cause of WS, and warrant further investigation of neurons and neuron-oligodendroglia interactions as a target for future neuroprotective or -restorative treatments for WS.

Deletion of the hypoxia-response element in the vascular endothelial growth factor promoter causes motor neuron degeneration.

Hypoxia stimulates angiogenesis through the binding of hypoxia-inducible factors to the hypoxia-response element in the vascular endothelial growth factor (Vegf) promotor. Here, we report that deletion of the hypoxia-response element in the Vegf promotor reduced hypoxic Vegf expression in the spinal cord and caused adult-onset progressive motor neuron degeneration, reminiscent of amyotrophic lateral sclerosis. The neurodegeneration seemed to be due to reduced neural vascular perfusion. In addition, Vegf165 promoted survival of motor neurons during hypoxia through binding to Vegf receptor 2 and neuropilin 1. Acute ischemia is known to cause nonselective neuronal death. Our results indicate that chronic vascular insufficiency and, possibly, insufficient Vegf-dependent neuroprotection lead to the select degeneration of motor neurons.

[Short-term dialectical behaviour therapy for borderline personality disorder]. / Kortdurende klinische dialectische gedragstherapie voor de borderlinepersoonlijkheidsstoornis: ontwerp van programma en resultaten pilotstudie.

BACKGROUND: Outpatient dialectical behaviour therapy (DBT) reduces severe suicidal and self-injurious behaviour in patients with borderline personality disorder. The Jelgersma center for personality disorders has developed an intensive inpatient dbt programme that lasts for 14 weeks and is designed to achieve a faster reduction in these borderline symptoms. AIM: To examine the effect of the Jelgersma programme by means of a pilot study in order to prepare a randomised clinical trial in which a short intensive course of DBT will be compared with standard outpatient DBT. METHOD: We compared the starting data and the final data for 39 female patients with borderline problems (DBT). We participated in 3½-month-long inpatient DBT programme. The collected data referred to (para)suicidal behaviour, drop-out, severity of borderline problems and the quality of life. RESULTS: The severity of borderline problems, particularly in the field of interpersonal problems, was significantly reduced. There was no significant reduction in (para) suicidal behaviours. The drop-out percentage was higher than in comparable studies. CONCLUSION: Short-term inpatient DBT had a positive effect on borderline problems. (Para)suicidal behaviour, however, was not reduced significantly. The randomised trial that began in 2012 should reveal whether the use of short-term inpatient DBT can lead to a faster decline of suicidal and self-injurious behaviour than does standard outpatient DBT.

Dialectical behavior therapy: is outpatient group psychotherapy an effective alternative to individual psychotherapy?: Preliminary conclusions.

OBJECTIVES: This study evaluates a 12-month-duration adapted outpatient group dialectical behavior therapy (DBT) program for patients with a borderline personality disorder in an unselected, comorbid population. If the results of this approach are comparable with the outcome rates of a standard DBT program, the group approach can have several advantages over individual treatment. One advantage is the possibility of treating more patients per therapist. METHOD: A pre-post design was used to measure the effectiveness of an outpatient group DBT. Data from the Beck Depression Inventory II, the Symptom Checklist 90-Revised, the State-Trait Anger Inventory, the State and Trait Anxiety Inventory, of 34 female patients (mean age, 32.65 years) were collected before and after a treatment period of 1 year. RESULTS: Overall, a significant reduction (P < .05) of depressive symptoms, suicidal thoughts, anxiety, and anger was experienced by the patients. CONCLUSIONS: This study is a first attempt in showing that DBT in an outpatient group setting can be effective in reducing psychiatric complaints and therefore has several advantages, such as the opportunity to treat more patients at once.

Genetic rodent models of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the selective death of motor neurons in the motor cortex, brainstem, and spinal cord. A large number of rodent models are available that show motor neuron death and a progressive motor phenotype that is more or less reminiscent of what occurs in patients. These rodent models contain genes with spontaneous or induced mutations or (over) express different (mutant) genes. Some of these models have been of great value to delineate potential pathogenic mechanisms that cause and/or modulate selective motor neuron degeneration. In addition, these genetic rodent models play a crucial role in testing and selecting potential therapeutics that can be used to treat ALS and/or other motor neuron disorders. In this paper, we give a systematic overview of the most important genetic rodent models that show motor neuron degeneration and/or develop a motor phenotype. In addition, we discuss the value and limitations of the different models and conclude that it remains a challenge to find more and better rodent models based on mutations in new genes causing ALS.

Differential contribution of the Na(+)-K(+)-2Cl(-) cotransporter NKCC1 to chloride handling in rat embryonic dorsal root ganglion neurons and motor neurons.

Plasma membrane chloride (Cl(-)) pathways play an important role in neuronal physiology. Here, we investigated the role of NKCC1 cotransporters (a secondary active Cl(-) uptake mechanism) in Cl(-) handling in cultured rat dorsal root ganglion neurons (DRGNs) and motor neurons (MNs) derived from fetal stage embryonic day 14. Gramicidin-perforated patch-clamp recordings revealed that DRGNs accumulate intracellular Cl(-) through a bumetanide- and Na(+)-sensitive mechanism, indicative of the functional expression of NKCC1. Western blotting confirmed the expression of NKCC1 in both DRGNs and MNs, but immunocytochemistry experiments showed a restricted expression in dendrites of MNs, which contrasts with a homogeneous expression in DRGNs. Both MNs and DRGNs could be readily loaded with or depleted of Cl(-) during GABA(A) receptor activation at depolarizing or hyperpolarizing membrane potentials. After loading, the rate of recovery to the resting Cl(-) concentration (i.e., [Cl(-)](i) decrease) was similar in both cell types and was unaffected by lowering the extracellular Na(+) concentration. In contrast, the recovery on depletion (i.e., [Cl(-)](i) increase) was significantly faster in DRGNs in control conditions but not in low extracellular Na(+). The experimental observations could be reproduced by a mathematical model for intracellular Cl(-) kinetics, in which DRGNs show higher NKCC1 activity and smaller Cl(-)-handling volume than MNs. On the basis of these results, we conclude that embryonic DRGNs show a higher somatic functional expression of NKCC1 than embryonic MNs. The high NKCC1 activity in DRGNs is important for maintaining high [Cl(-)](i), whereas lower NKCC1 activity in MNs allows large [Cl(-)](i) variations during neuronal activity.

[Dialectical behaviour therapy for adolescents; a literature review]. / Dialectische gedragstherapie bij adolescenten;een literatuuronderzoek.

BACKGROUND: According to several randomised controlled trials (rct's) dialectical behaviour therapy (dbt) is effective in treating adults diagnosed with borderline personality disorder (bpd) who present with self-injurious and suicidal behaviour. In recent years there have been several studies about dbt in adolescents with varying problems and disorders. AIM: To review the literature for evidence of the effectiveness of dbt in adolescents aged 12 to 18. METHOD: With the help of PubMed and Medline and using the search-terms 'dialectical', 'adolescent', 'suicide attempt' and 'deliberate self harm', we searched the literature for references to dbt in adolescents. RESULTS: There were no rct's involving dbt in adolescents, but we did find one quasi-experimental design and several other studies with a pre-post treatment design. However, the studies were difficult to compare. In some cases it was doubtful whether the treatment could still be called dbt. The results suggested that dbt may be just as effective with adolescents as it is with adults in reducing bpd symptoms, suicidal ideation, and comorbid depressive disorder symptoms, and in reducing the need for hospitalisation. The results also indicated that dbt might be effective in treating eating disorders, bipolar disorder, oppositionality, aggression and nonsuicidal self-injurious behaviour (nsib) in a variety of treatment settings. CONCLUSION: dbt is possibly effective for treating adolescents with nsib and/or bpd symptoms. It may also be an effective treatment for various other affective and behavioural disorders. rct's are needed.

Probabilistic risk assessment model for allergens in food: sensitivity analysis of the minimum eliciting dose and food consumption.

Previously, TNO developed a probabilistic model to predict the likelihood of an allergic reaction, resulting in a quantitative assessment of the risk associated with unintended exposure to food allergens. The likelihood is estimated by including in the model the proportion of the population who is allergic, the proportion consuming the food and the amount consumed, the likelihood of the food containing an adventitious allergen and its concentration, and the minimum eliciting dose (MED) distribution for the allergen. In the present work a sensitivity analysis was performed to identify which parts of the model most influence the output. A shift in the distribution of the MED reflecting a more potent allergen, and an increase in the proportion of the population consuming a food, increased the number of estimated allergic reactions considerably. In contrast, the number of estimated allergic reactions hardly changed when the MEDs were based on a more severe response, or when the amount of food consumed was increased. Development of this work will help to generate a more accurate picture of the potential public health impact of allergens. It highlights areas where research is best focused, specifically the determination of minimum eliciting doses and understanding of the food choices of allergic individuals.

Treatment of antisocial personality disorder: Development of a practice focused framework.

There is little to no evidence of effective treatment methods for patients with an antisocial personality disorder (ASPD). One of the reasons could be the fact that they are often excluded from mental healthcare and thus from studies. A treatment framework based on 'state of the art' methods and best practices, offering guidelines on the treatment of ASP and possibilities for more systematical research, is urgently needed. This research involved a literature search and an international Delphi-study (N = 61 experts in research, management and clinical practice focused on ASPD). The results suggested important preconditions with regard to organization of care, healthcare workers and therapy. Conclusions are that there are many ways to coordinate effective treatment and management and work toward the increased availability of evidence based care for persons with ASPD.

Integrated molecular landscape of amyotrophic lateral sclerosis provides insights into disease etiology.

Amyotrophic lateral sclerosis (ALS) is a severe, progressive and ultimately fatal motor neuron disease caused by a combination of genetic and environmental factors, but its underlying mechanisms are largely unknown. To gain insight into the etiology of ALS, we here conducted genetic network and literature analyses of the top-ranked findings from six genome-wide association studies of sporadic ALS (involving 3589 cases and 8577 controls) as well as genes implicated in ALS etiology through other evidence, including familial ALS candidate gene association studies. We integrated these findings into a molecular landscape of ALS that allowed the identification of three main processes that interact with each other and are crucial to maintain axonal functionality, especially of the long axons of motor neurons, i.e. (1) Rho-GTPase signaling; (2) signaling involving the three regulatory molecules estradiol, folate, and methionine; and (3) ribonucleoprotein granule functioning and axonal transport. Interestingly, estradiol signaling is functionally involved in all three cascades and as such an important mediator of the molecular ALS landscape. Furthermore, epidemiological findings together with an analysis of possible gender effects in our own cohort of sporadic ALS patients indicated that estradiol may be a protective factor, especially for bulbar-onset ALS. Taken together, our molecular landscape of ALS suggests that abnormalities within three interconnected molecular processes involved in the functioning and maintenance of motor neuron axons are important in the etiology of ALS. Moreover, estradiol appears to be an important modulator of the ALS landscape, providing important clues for the development of novel disease-modifying treatments.

The role of excitotoxicity in the pathogenesis of amyotrophic lateral sclerosis.

Unfortunately and despite all efforts, amyotrophic lateral sclerosis (ALS) remains an incurable neurodegenerative disorder characterized by the progressive and selective death of motor neurons. The cause of this process is mostly unknown, but evidence is available that excitotoxicity plays an important role. In this review, we will give an overview of the arguments in favor of the involvement of excitotoxicity in ALS. The most important one is that the only drug proven to slow the disease process in humans, riluzole, has anti-excitotoxic properties. Moreover, consumption of excitotoxins can give rise to selective motor neuron death, indicating that motor neurons are extremely sensitive to excessive stimulation of glutamate receptors. We will summarize the intrinsic properties of motor neurons that could render these cells particularly sensitive to excitotoxicity. Most of these characteristics relate to the way motor neurons handle Ca(2+), as they combine two exceptional characteristics: a low Ca(2+)-buffering capacity and a high number of Ca(2+)-permeable AMPA receptors. These properties most likely are essential to perform their normal function, but under pathological conditions they could become responsible for the selective death of motor neurons. In order to achieve this worst-case scenario, additional factors/mechanisms could be required. In 1 to 2% of the ALS patients, mutations in the SOD1 gene could shift the balance from normal motor neuron excitation to excitotoxicity by decreasing glutamate uptake in the surrounding astrocytes and/or by interfering with mitochondrial function. We will discuss point by point these different pathogenic mechanisms that could give rise to classical and/or slow excitotoxicity leading to selective motor neuron death.

[The causes and mechanism of selective motor neuron death in amyotrophic lateral sclerosis]. / Onderzoek naar de oorzaken en het mechanisme van de selectieve motorneurondood bij Amyotrofische Lateraal Sclerose (ALS).

Amyotrophic lateral sclerosis is a neurodegenerative disease characterized by the selective death of motor neurons in the motor cortex, brain stem and spinal cord. In most cases, the cause of ALS is unknown although in a number of familial ALS cases mutations in the superoxide dismutase 1 (SOD1) gene were discovered. The mechanism underlying the selective motor neuron death is not yet clarified. However, it is clear that excitotoxicity could play a role in the selectivity of the motor neuron death. Excitotoxicity is the phenomenon in which the normal glutamate-mediated communication between neurons is disturbed and ultimately leads to neuronal death. In the first part of this study, we have investigated the intrinsic characteristics of motor neurons that could be responsible for the fact that these cells are extremely sensitive to excitotoxicity. Despite the fact that only the motor neurons die during ALS, it is clear that also other cell types play an important role during this process. In the second part of this study, we have concentrated on the potential role of the microglia. These cells are the macrophages of the brain and they become activated during inflammation. Using minocycline, we have inhibited the activation of the microglia and we have investigated its effect on the start of the disease and on the survival of a mouse model for ALS. Furthermore, we have investigated the cellular mechanism underlying the effect of minocycline. The goal of this research is to get insight into the mechanisms responsible for the selective motor neuron death during ALS and we hope that this information can help in the development of a therapy for this dramatic and incurable disease.

Chloride influx aggravates Ca2+-dependent AMPA receptor-mediated motoneuron death.

AMPA receptor-mediated excitotoxicity has been implicated in the pathogenesis of stroke, neurotrauma, epilepsy, and many neurodegenerative diseases such as motoneuron disease. We studied the role of Cl- in AMPA receptor-mediated Ca2+-dependent excitotoxicity in cultured rat spinal motoneurons. Using the gramicidin perforated patch-clamp technique, the intracellular Cl- concentration could be calculated from the reversal potential of the GABA-induced current. The membrane depolarization caused by AMPA receptor stimulation resulted in Cl- influx through 5-nitro-2(3-phenylpropyl-amino) benzoic acid- and niflumic acid-sensitive Cl- channels. Cl- influx during AMPA receptor stimulation aggravated excitotoxic motoneuron death by two mechanisms: an increase of AMPA receptor conductance and an elevation of the Ca2+ driving force through a partial repolarization. The Cl- influx during AMPA receptor stimulation was enhanced by coadministration of GABA. This resulted in an increased Ca2+ influx and an enhanced cell death, suggesting that concomitant GABAergic stimulation may aggravate excitotoxic motoneuron death.

Polyamine transport systems in the LLC-PK1 renal epithelial established cell line.

LLC-PK1 cells were brought to a quiescent state by treatment with DL-2-difluoromethylornithine (DFMO), a specific inhibitor of L-ornithine decarboxylase (ODC). The inhibition of ODC, which is the key enzyme for polyamine synthesis, strongly reduced the cellular content of putrescine and spermidine. The cells resumed DNA-synthesis followed by mitosis when exogenous putrescine was added. DFMO treatment strongly stimulated the putrescine uptake capability. A kinetic analysis of the initial uptake rates revealed a saturable Na+-dependent and a saturable Na+-independent pathway on top of non-saturable diffusion. The stimulation by DFMO was exclusively due to an effect on the Vmax values of the saturable pathways. The Na+-dependent transporter had a higher affinity for putrescine (apparent Km = 4.7 +/- 0.7 microM) than the Na+-independent transporter (apparent Km = 29.8 +/- 3.5 microM). As a consequence, although the latter transporter had a higher Vmax, the Na+-dependent transport was more important at a physiological putrescine concentration. Putrescine uptake by both transporters was inhibited with similar relative affinities by spermidine, spermine as well as by the antileukemic agent, methylglyoxal bis(guanylhydrazone), but not by amino acids. The activity of the Na+-dependent transporter was very much dependent on SH-group reagents, whereas the Na+-independent transporter was not affected. Both transporters were inhibited by metabolic inhibitors and by ionophores but the Na+-dependent transporter was affected to a greater extent. For both transporters there was a down-regulation in response to exogenous putrescine. This suggests that the polyamine transporters in LLC-PK1 are adaptively regulated and may contribute to the regulation of the cellular polyamine level and cellular proliferation.