GLRB allelic variation associated with agoraphobic cognitions, increased startle response and fear network activation : a potential neurogenetic pathway to panic disorder

The molecular genetics of panic disorder (PD) with and without agoraphobia (AG) are still largely unknown and progress is hampered by small sample sizes. We therefore performed a genome-wide association study with a dimensional, PD/AG - related anxiety phenotype based on the Agoraphobia Cognition Questionnaire (ACQ) in a sample of 1,370 healthy German volunteers of the CRC TRR58 MEGA study wave 1. A genome-wide significant association was found between ACQ and single non-coding nucleotide variants of the GLRB gene (rs78726293, p=3.3x10-8; rs191260602, p=3.9x10-8). We followed up on this finding in a larger dimensional ACQ sample (N=2,547) and in independent samples with a dichotomous AG phenotype based on the Symptoms Checklist (SCL-90; N=3,845) and a case control sample with the categorical phenotype PD/AG (Ncombined =1,012) obtaining highly significant p-values also for GLRB single nucleotide variants rs17035816 (p=3.8x10-4) and rs7688285 (p=7.6x10-5). GLRB gene expression was found to be modulated by rs7688285 in brain tissue as well as cell culture. Analyses of intermediate PD/AG phenotypes demonstrated increased startle reflex and increased fear network as well as general sensory activation by GLRB risk gene variants rs78726293, rs191260602, rs17035816 and rs7688285. Partial Glrb knockout-mice demonstrated an agoraphobic phenotype. In conjunction withthe clinical observation that rare coding GLRB gene mutations are associated with the neurological disorder hyperekplexia characterized by a generalized startle reaction and agoraphobic behavior, our data provide evidence that non-coding, though functional GLRB gene polymorphisms may predispose to PD by increasing startle response and agoraphobic cognitions.PostprintPeer reviewe

Astrozytenproliferation in Ratten und Mäusen – Hinweise für eine regulierende Rolle von Dopamine und Aquaporin 4

Astrocytes of the striatum express both dopamine receptors and transporters, which indicates that dopamine plays a functional role in striatal astrocytes. Dopamine is known to modulate proliferation of precursor cells during development and also in the adult brain. However, it is unknown whether dopamine affects proliferation of astrocytes. In this study we investigated a putative role of dopamine on the proliferation of striatal astrocytes in vitro and in vivo. Using striatal mouse astrocyte cultures we found that dopamine decreased proliferation of astrocytes and the expression of the water channel AQP4, more precisely the isoform M23, both on the mRNA and protein levels. We hypothesized that the decreased expression of the water channel is linked to the observed decrease in proliferation. Under physiological conditions, the striatum represents a target area of dopaminergic projections arising in the midbrain. We hypothesized that dopamine might keep AQP4 expression at a low level and hence prevent proliferation of astrocytes in the striatum in vivo. Consequentially, in vivo depletion of dopamine should result in an increased expression of AQP4 and an increase of proliferation of astrocytes. To investigate this hypothesis we intraventricularily injected 6-hydroxydopamine, a catecholaminergic neurotoxin that destroys dopaminergic neurons. Under these conditions AQP4 mRNA expression, specifically the AQP4 M23 isoform, was increased in the striatum, and we observed that proliferation was significantly increased in the striatum and the lateral cortex, both regions that were affected by the loss of dopamine.Astrozyten im Striatum exprimieren Dopaminerezeptoren und Transporter, was darauf hinweisst dass Dopamine eine regulierende Funtion in striatalen Astrozyten besitzt. Es ist gezeigt, dass Dopamine die Proliferation von Vorläuferzellen während der Entwicklung moduliert, jedoch ist nicht bekannt ob Dopamin auch die Proliferation von Astrozyten beeinflusst. In dieser Studie wurde die mögliche Rolle von Dopamin auf die Astrozytenproliferation untersucht, jeweils in vitro und in vivo. In striatalen Mäuseastrozytenkulturen beobachteten wir nach Behandlung der Zellen mit Dopamin eine verminderte Proliferation und eine verringerte Expression des Wasserkanals Aquaporin 4, insbesondere die Isoform M23. Wir vermuteten, dass die verringerte Expression des Wasserkanals mit der verminderten Proliferation zusammenhängt. Im gesunden Gehirn ist das Striatum eine stark dopaminerg-innervierte Region. Wenn unsere erste Vermutung stimmt, müsste also das Vorhandensein von Dopamin die Aquaporin 4 Expression auf einen niedrigen Level halten und damit die Proliferation von Astrozyten verhindern. Folglich, sollte es zu einem Verlust von Dopamin im Striatum kommen, müsste man eine erhöhte Astrozytenproliferation und AQP4 Expression finden. Um diese Hypothese zu untersuchen, injizierten wir das Neurotoxin 6-Hydroxydopamin, welches selektiv für catecholaminerge Neurone ist, in die lateralen Ventrikel. Unter diesen Bedingungen beobachteten wir im Striatum eine erhöhte AQP4 Expression, vor allem M23, sowie einen signifikanten Anstieg an proliferierenden Astrozyten

Anxiety and Startle Phenotypes in Glrb Spastic and Glra1 Spasmodic Mouse Mutants

A GWAS study recently demonstrated single nucleotide polymorphisms (SNPs) in the human GLRB gene of individuals with a prevalence for agoraphobia. GLRB encodes the glycine receptor (GlyRs) β subunit. The identified SNPs are localized within the gene flanking regions (3′ and 5′ UTRs) and intronic regions. It was suggested that these nucleotide polymorphisms modify GlyRs expression and phenotypic behavior in humans contributing to an anxiety phenotype as a mild form of hyperekplexia. Hyperekplexia is a human neuromotor disorder with massive startle phenotypes due to mutations in genes encoding GlyRs subunits. GLRA1 mutations have been more commonly observed than GLRB mutations. If an anxiety phenotype contributes to the hyperekplexia disease pattern has not been investigated yet. Here, we compared two mouse models harboring either a mutation in the murine Glra1 or Glrb gene with regard to anxiety and startle phenotypes. Homozygous spasmodic animals carrying a Glra1 point mutation (alanine 52 to serine) displayed abnormally enhanced startle responses. Moreover, spasmodic mice exhibited significant changes in fear-related behaviors (freezing, rearing and time spent on back) analyzed during the startle paradigm, even in a neutral context. Spastic mice exhibit reduced expression levels of the full-length GlyRs β subunit due to aberrant splicing of the Glrb gene. Heterozygous animals appear normal without an obvious behavioral phenotype and thus might reflect the human situation analyzed in the GWAS study on agoraphobia and startle. In contrast to spasmodic mice, heterozygous spastic animals revealed no startle phenotype in a neutral as well as a conditioning context. Other mechanisms such as a modulatory function of the GlyRs β subunit within glycinergic circuits in neuronal networks important for fear and fear-related behavior may exist. Possibly, in human additional changes in fear and fear-related circuits either due to gene-gene interactions e.g., with GLRA1 genes or epigenetic factors are necessary to create the agoraphobia and in particular the startle phenotype