Deep Survey of GABAergic Interneurons: Emerging Insights From Gene-Isoform Transcriptomics

GABAergic interneuron diversity is a key feature in the brain that helps to create different brain activity patterns and behavioral states. Cell type classification schemes—based on anatomical, physiological and molecular features—have provided us with a detailed understanding of the distinct types that constitute this diversity and their contribution to brain function. Over recent years, the utility of single-cell RNAseq has majorly complemented this existing framework, vastly expanding our knowledge base, particularly regarding molecular features. Single-cell gene-expression profiles of tens of thousands of GABAergic cells from many different types are now available. The analysis of these data has shed new lights onto previous classification principles and illuminates a path towards a deeper understanding of molecular hallmarks behind interneuron diversity. A large part of such molecular features is synapse-related. These include ion channels and receptors, as well as key synaptic organizers and trans-synaptic signaling molecules. Increasing evidence suggests that transcriptional and post-transcriptional modifications further diversify these molecules and generate cell type-specific features. Thus, unraveling the cell type-specific nature of gene-isoform expression will be a key in cell type classification. This review article discusses progress in the transcriptomic survey of interneurons and insights that have begun to manifest from isoform-level analyses

Single-Cell RNA-Seq Reveals Developmental Origins and Ontogenetic Stability of Neurexin Alternative Splicing Profiles

Neurexins are key synaptic organizers that are expressed in thousands of alternatively spliced isoforms. Because transsynaptic neurexin interactions with different postsynaptic molecules are largely isoform dependent, a cell type-level census of different neurexin isoforms could predict molecular interactions relating to synapse identity and function. Using single-cell transcriptomics to study the origin of neurexin diversity in multiple murine mature and embryonic cell types, we have discovered shared neurexin expression patterns in developmentally related cells. By comparing neurexin profiles in immature embryonic neurons, we show that neurexin profiles are specified during early development and remain unchanged throughout neuronal maturation. Thus, our findings reveal ontogenetic stability and provide a cell type-level census of neurexin isoform expression in the cortex

Transcriptomically-guided pharmacological experiments in neocortical and hippocampal NPY positive GABAergic interneurons

Cortical GABAergic interneurons have been shown to fulfil important roles by inhibiting excitatory principal neurons. Recent transcriptomic studies have confirmed seminal discoveries that used anatomic and electrophysiological methods highlighting the existence of multiple different classes of GABAergic interneurons. Although some of these studies have emphasized that inter-regional differences may exist for a given class, the extent of such differences remains unknown. To address this problem, we used single-cell Patch-RNAseq to characterize neuropeptide Y (NPY)-positive GABAergic interneurons in superficial layers of the primary auditory cortex (AC) and in distal layers of area CA3 in mice. We found that more than 300 genes are differentially expressed in NPY-positive neurons between these two brain regions. For example, the AMPA receptor (AMPAR) auxiliary subunit Shisa9/CKAMP44 and the 5HT2a receptor (5HT2aR) are significantly higher expressed in auditory NPY-positive neurons. These findings guided us to perform pharmacological experiments that revealed a role for 5HT2aRs in auditory NPY-positive neurons. Specifically, although the application of 5HT led to a depolarization of both auditory and CA3 NPY-positive neurons, the 5HT2aR antagonist ketanserin only reversed membrane potential changes in auditory NPY-positive neurons. Our study demonstrates the potential of single-cell transcriptomic studies in guiding directed pharmacological experiments

Excitatory microcircuits within superficial layers of the medial entorhinal cortex

The distinctive firing pattern of grid cells in the medial entorhinal cortex (MEC) supports its role in the representation of space. It is widely believed that the hexagonal firing field of grid cells emerges from neural dynamics that depends on the local microcircuitry. However, local networks within the MEC are still not sufficiently characterized. Here, applying up to eight simultaneous whole-cell recordings in acute brain slices, we demonstrate the existence of unitary excitatory connections between principal neurons in the superficial layers of the MEC. In particular, we find prevalent feed-forward excitation from pyramidal neurons in layer III and layer II onto stellate cells in layer II, which might contribute to the generation or the inheritance of grid-cell patterns

Preoperative medication use and development of postoperative delirium and cognitive dysfunction

Postoperative delirium (POD) and postoperative (neuro-)cognitive disorder (POCD) are frequent and serious complications after operations. We aim to investigate the association between pre-operative polypharmacy and potentially inappropriate medications and the development of POD/POCD in elderly patients. This investigation is part of the European BioCog project (www.biocog.eu), a prospective multicenter observational study with elderly surgical patients. Patients with a Mini-Mental State Examination score less than or equal to 23 points were excluded. POD was assessed up to 7 days after surgery using the Nursing Delirium Screening Scale, Confusion Assessment Method (for the intensive care unit [ICU]), and a patient chart review. POCD was assessed 3 months after surgery with a neuropsychological test battery. Pre-operative long-term medication was evaluated in terms of polypharmacy (≥5 agents) and potentially inappropriate medication (defined by the PRISCUS and European list of potentially inappropriate medications [EU(7)-PIM] lists), and associations with POD and POCD were analyzed using logistic regression analysis. Eight hundred thirty-seven participants were included for analysis of POD and 562 participants for POCD. Of these, 165 patients (19.7%) fulfilled the criteria of POD and 60 (10.7%) for POCD. After adjusting for confounders, pre-operative polypharmacy and intake of potentially inappropriate medications could not be shown to be associated with the development of POD nor POCD. We found no associations between pre-operative polypharmacy and potentially inappropriate medications and development of POD and POCD. Future studies should focus on the evaluation of drug interactions to determine whether patients benefit from a pre-operative adjustment

Commissural dentate granule cell projections and their rapid formation in the adult brain

Dentate granule cells (GCs) have been characterized as unilaterally projecting neurons within each hippocampus. Here, we describe a unique class, the commissural GCs, which atypically project to the contralateral hippocampus in mice. Although commissural GCs are rare in the healthy brain, their number and contralateral axon density rapidly increase in a rodent model of temporal lobe epilepsies. In this model, commissural GC axon growth appears together with the well-studied hippocampal mossy fiber sprouting and may be important for the pathomechanisms of epilepsy. Our results augment the current view on hippocampal GC diversity and demonstrate powerful activation of a commissural wiring program in the adult brain

Functional specification of CCK plus interneurons by alternative isoforms of Kv4.3 auxiliary subunits

CCK-expressing interneurons (CCK+INs) are crucial for controlling hippocampal activity. We found two firing phenotypes of CCK+INs in rat hippocampal CA3 area; either possessing a previously undetected membrane potential-dependent firing or regular firing phenotype, due to different low-voltage-activated potassium currents. These different excitability properties destine the two types for distinct functions, because the former is essentially silenced during realistic 8-15 Hz oscillations. By contrast, the general intrinsic excitability, morphology and gene-profiles of the two types were surprisingly similar. Even the expression of Kv4.3 channels were comparable, despite evidences showing that Kv4.3-mediated currents underlie the distinct firing properties. Instead, the firing phenotypes were correlated with the presence of distinct isoforms of Kv4 auxiliary subunits (KChIP1 vs. KChIP4e and DPP6S). Our results reveal the underlying mechanisms of two previously unknown types of CCK+INs and demonstrate that alternative splicing of few genes, which may be viewed as a minor change in the cells' whole transcriptome, can determine cell-type identity

Preoperative comparison of three anticholinergic drug scales in older adult patients and development of postoperative delirium: a prospective observational study

BACKGROUND: Postoperative delirium (POD) is a frequent and serious complication after surgery. Evidence of a relationship between anticholinergic medication and the development of delirium is inconclusive, but studies on POD are rare. OBJECTIVES: The objective of this study was to evaluate the anticholinergic load of preoperative medication in older adult patients and its association with the development of POD. METHODS: This investigation was part of the European BioCog project ( http://www.biocog.eu ), a prospective multicenter observational study in older adult surgical patients (ClinicalTrials.gov identifier: NCT02265263, 15 October 2014). Patients with a Mini-Mental State Examination score ≤ 23 points were excluded. POD was assessed up to 7 days after surgery using the Nursing Delirium Screening Scale, Confusion Assessment Method and a patient chart review. The preoperative anticholinergic load was calculated using the Anticholinergic Drug Scale (ADS), the Anticholinergic Risk Scale (ARS) and the Anticholinergic Cognitive Burden Scale (ACBS), and associations with POD were analyzed using logistic regression analysis adjusting for age, comorbidities, duration of anesthesia and number of drugs used. RESULTS: In total, 837 participants were included for analysis, and 165 patients (19.7%) fulfilled the criteria of POD. After adjusting for confounders, we found no association between preoperative anticholinergic load and the development of POD (ADS [points] odds ratio [OR] 0.928; 95% confidence interval [CI] 0.749-1.150; ARS [points] OR 0.832; 95% CI 0.564-1.227; ACBS [points] OR 1.045; 95% CI 0.842-1.296). CONCLUSION: This study found no association between the anticholinergic load of drugs used preoperatively and the development of POD in older adult patients without severe preexisting cognitive impairment. Future analyses should examine the influence of intra- and postoperative administration of anticholinergic drugs as well as dosages of and interactions between medications

Preoperative medication use and development of postoperative delirium and cognitive dysfunction

Postoperative delirium (POD) and postoperative (neuro-)cognitive disorder (POCD) are frequent and serious complications after operations. We aim to investigate the association between pre-operative polypharmacy and potentially inappropriate medications and the development of POD/POCD in elderly patients. This investigation is part of the European BioCog project (www.biocog.eu), a prospective multicenter observational study with elderly surgical patients. Patients with a Mini-Mental State Examination score less than or equal to 23 points were excluded. POD was assessed up to 7 days after surgery using the Nursing Delirium Screening Scale, Confusion Assessment Method (for the intensive care unit [ICU]), and a patient chart review. POCD was assessed 3 months after surgery with a neuropsychological test battery. Pre-operative long-term medication was evaluated in terms of polypharmacy (≥5 agents) and potentially inappropriate medication (defined by the PRISCUS and European list of potentially inappropriate medications [EU(7)-PIM] lists), and associations with POD and POCD were analyzed using logistic regression analysis. Eight hundred thirty-seven participants were included for analysis of POD and 562 participants for POCD. Of these, 165 patients (19.7%) fulfilled the criteria of POD and 60 (10.7%) for POCD. After adjusting for confounders, pre-operative polypharmacy and intake of potentially inappropriate medications could not be shown to be associated with the development of POD nor POCD. We found no associations between pre-operative polypharmacy and potentially inappropriate medications and development of POD and POCD. Future studies should focus on the evaluation of drug interactions to determine whether patients benefit from a pre-operative adjustment

Functional polymorphism in the neuropeptide Y gene promoter (rs16147) is associated with serum leptin levels and waist-hip ratio in women

OBJECTIVE: The neuropeptide-Y (NP-Y) gene is a strong candidate gene in the pathophysiology of obesity-linked behavior, and several single-nucleotide polymorphisms of NP-Y have already been linked to body weight and appetite. However, the results from current studies remain inconclusive. The aim of the present study was to test whether a certain functional genetic variant (SNP rs16147) in the NP-Y promoter gene is associated with serum leptin levels and body fat distribution. METHOD: We genotyped and measured the serum leptin levels of the NP-Y rs16147 polymorphism in 1,097 Caucasian subjects in the context of a population-based, case-control multicenter study. We measured weight, height and waist circumference, from which we then calculated BMI and waist-to-hip ratio (WHR). RESULTS: We found the CT-genotype of the SNP rs16147 to be significantly associated with lower WHRs and higher serum leptin levels in women, compared to homozygote gene carriers. No association between rs16147, WHR and serum leptin levels was found in men. CONCLUSION: Our results provide evidence that the functionally relevant SNP in the NP-Y promoter gene affects body fat distribution and serum leptin levels in women, pointing towards possible behavioral effects of NPY in obesity