Altered neurological and neurobehavioral phenotypes in a mouse model of the recurrent KCNB1-p.R306C voltage-sensor variant.

Pathogenic variants in KCNB1 are associated with a neurodevelopmental disorder spectrum that includes global developmental delays, cognitive impairment, abnormal electroencephalogram (EEG) patterns, and epilepsy with variable age of onset and severity. Additionally, there are prominent behavioral disturbances, including hyperactivity, aggression, and features of autism spectrum disorder. The most frequently identified recurrent variant is KCNB1-p.R306C, a missense variant located within the S4 voltage-sensing transmembrane domain. Individuals with the R306C variant exhibit mild to severe developmental delays, behavioral disorders, and a diverse spectrum of seizures. Previous in vitro characterization of R306C described altered sensitivity and cooperativity of the voltage sensor and impaired capacity for repetitive firing of neurons. Existing Kcnb1 mouse models include dominant negative missense variants, as well as knockout and frameshifts alleles. While all models recapitulate key features of KCNB1 encephalopathy, mice with dominant negative alleles were more severely affected. In contrast to existing loss-of-function and dominant-negative variants, KCNB1-p.R306C does not affect channel expression, but rather affects voltage-sensing. Thus, modeling R306C in mice provides a novel opportunity to explore impacts of a voltage-sensing mutation in Kcnb1. Using CRISPR/Cas9 genome editing, we generated the Kcnb1R306C mouse model and characterized the molecular and phenotypic effects. Consistent with the in vitro studies, neurons from Kcnb1R306C mice showed altered excitability. Heterozygous and homozygous R306C mice exhibited hyperactivity, altered susceptibility to chemoconvulsant-induced seizures, and frequent, long runs of slow spike wave discharges on EEG, reminiscent of the slow spike and wave activity characteristic of Lennox Gastaut syndrome. This novel model of channel dysfunction in Kcnb1 provides an additional, valuable tool to study KCNB1 encephalopathies. Furthermore, this allelic series of Kcnb1 mouse models will provide a unique platform to evaluate targeted therapies.

Altered neurological and neurobehavioral phenotypes in a mouse model of the recurrent KCNB1-p.R306C voltage-sensor variant.

Pathogenic variants in KCNB1 are associated with a neurodevelopmental disorder spectrum that includes global developmental delays, cognitive impairment, abnormal electroencephalogram (EEG) patterns, and epilepsy with variable age of onset and severity. Additionally, there are prominent behavioral disturbances, including hyperactivity, aggression, and features of autism spectrum disorder. The most frequently identified recurrent variant is KCNB1-p.R306C, a missense variant located within the S4 voltage-sensing transmembrane domain. Individuals with the R306C variant exhibit mild to severe developmental delays, behavioral disorders, and a diverse spectrum of seizures. Previous in vitro characterization of R306C described loss of voltage sensitivity and cooperativity of the sensor and inhibition of repetitive firing. Existing Kcnb1 mouse models include dominant negative missense variants, as well as knockout and frameshifts alleles. While all models recapitulate key features of KCNB1 encephalopathy, mice with dominant negative alleles were more severely affected. In contrast to existing loss-of-function and dominant-negative variants, KCNB1-p.R306C does not affect channel expression, but rather affects voltage-sensing. Thus, modeling R306C in mice provides a novel opportunity to explore impacts of a voltage-sensing mutation in Kcnb1. Using CRISPR/Cas9 genome editing, we generated the Kcnb1R306C mouse model and characterized the molecular and phenotypic effects. Heterozygous and homozygous R306C mice exhibited pronounced hyperactivity, altered susceptibility to flurothyl and kainic acid induced-seizures, and frequent, long runs of spike wave discharges on EEG. This novel model of channel dysfunction in Kcnb1 provides an additional, valuable tool to study KCNB1 encephalopathies. Furthermore, this allelic series of Kcnb1 mouse models will provide a unique platform to evaluate targeted therapies.

Behavior and Regional Cortical BOLD Signal Fluctuations Are Altered in Adult Rabbits After Neonatal Volatile Anesthetic Exposure.

Neonatal and infant exposure to volatile anesthetics has been associated with long-term learning, memory, and behavioral deficits. Although early anesthesia exposure has been linked to a number of underlying structural abnormalities, functional changes associated with these impairments remain poorly understood. To investigate the relationship between functional alteration in neuronal circuits and learning deficiency, resting state functional MRI (rsfMRI) connectivity was examined in adolescent rabbits exposed to general anesthesia as neonates (1 MAC isoflurane for 2 h on postnatal days P8, P11, and P14) and unanesthetized controls before and after training with a trace eyeblink classical conditioning (ECC) paradigm. Long-range connectivity was measured between several key regions of interest (ROIs), including primary and secondary somatosensory cortices, thalamus, hippocampus, and cingulate. In addition, metrics of regional BOLD fluctuation amplitudes and coherence, amplitude of low-frequency fluctuation (ALFF), fractional ALFF (fALFF), and regional homogeneity (ReHo) were calculated. Our results showed that the trace ECC learning rate was significantly lower in the anesthesia-exposed group. No anesthesia-related changes in long-range connectivity, fALFF, or ReHo were found between any ROIs. However, ALFF was significantly higher in anesthesia-exposed rabbits in the primary and secondary somatosensory cortices, and ALFF in those areas was a significant predictor of the learning performance for trace ECC. The absence of anesthesia-related changes in long-range thalamocortical connectivity indicates that functional thalamocortical input is not affected. Higher ALFF in the somatosensory cortex may indicate the developmental disruption of cortical neuronal circuits after neonatal anesthesia exposure, including excessive neuronal synchronization that may underlie the observed cognitive deficits.

Effects of neonatal isoflurane anesthesia exposure on learning-specific and sensory systems in adults.

Millions of children undergo general anesthesia each year, and animal and human studies have indicated that exposure to anesthesia at an early age can impact neuronal development, leading to behavioral and learning impairments that manifest later in childhood and adolescence. Here, we examined the effects of isoflurane, a commonly-used general anesthetic, which was delivered to newborn rabbits. Trace eyeblink classical conditioning was used to assess the impact of neonatal anesthesia exposure on behavioral learning in adolescent subjects, and a variety of MRI techniques including fMRI, MR volumetry, spectroscopy and DTI captured functional, metabolic, and structural changes in key regions of the learning and sensory systems associated with anesthesia-induced learning impairment. Our results demonstrated a wide array of changes that were specific to anesthesia-exposed subjects, which supports previous studies that have pointed to a link between early anesthesia exposure and the development of learning and behavioral deficiencies. These findings point to the need for caution in avoiding excessive use of general anesthesia in young children and neonates.

Impact of anesthesia exposure in early development on learning and sensory functions.

Each year, millions of children undergo anesthesia, and both human and animal studies have indicated that exposure to anesthesia at an early age can lead to neuronal damage and learning deficiency. However, disorders of sensory functions were not reported in children or animals exposed to anesthesia during infancy, which is surprising, given the significant amount of damage to brain tissue reported in many animal studies. In this review, we discuss the relationship between the systems in the brain that mediate sensory input, spatial learning, and classical conditioning, and how these systems could be affected during anesthesia exposure. Based on previous reports, we conclude that anesthesia can induce structural, functional, and compensatory changes in both sensory and learning systems. Changes in myelination following anesthesia exposure were observed as well as the neurodegeneration in the gray matter across variety of brain regions. Disproportionate cell death between excitatory and inhibitory cells induced by anesthesia exposure can lead to a long-term shift in the excitatory/inhibitory balance, which affects both learning-specific networks and sensory systems. Anesthesia may directly affect synaptic plasticity which is especially critical to learning acquisition. However, sensory systems appear to have better ability to compensate for damage than learning-specific networks.

The effect of sevoflurane and isoflurane anesthesia on single unit and local field potentials.

Volatile general anesthetics are used commonly in adults and children, yet their mechanisms of action are complex and the changes in single unit firing and synaptic activity that underlie the broad decreases in neuronal activity induced by these drugs have not been well characterized. Capturing such changes throughout the anesthesia process is important for comparing the effects of different anesthetics and gaining a better understanding of their mechanisms of action and their impact on different brain regions. Using chronically implanted electrodes in the rabbit somatosensory cortex, we compared the effects of two common general anesthetics, isoflurane, and sevoflurane, on cortical neurons. Single unit activity and local field potentials (LFP) were recorded continuously before and during anesthetic delivery at 1 MAC, as well as during recovery. Our findings show that although isoflurane and sevoflurane belong to the same class of volatile general anesthetics, their effects upon cortical single units and LFP were quite different. Overall, the suppression of neuronal firing was greater and more uniform under sevoflurane. Moreover, the changes in LFP frequency bands suggest that effect of anesthesia upon beta oscillations does not necessarily depend on the level of single unit activity, but rather on the changes in GABA/glutamate neurotransmission induced by each drug.