Mechanisms Underlying the Rapid Induction and Sustained Expression of Synaptic Homeostasis

SummaryHomeostatic signaling systems are thought to interface with the mechanisms of neural plasticity to achieve stable yet flexible neural circuitry. However, the time course, molecular design, and implementation of homeostatic signaling remain poorly defined. Here we demonstrate that a homeostatic increase in presynaptic neurotransmitter release can be induced within minutes following postsynaptic glutamate receptor blockade. The rapid induction of synaptic homeostasis is independent of new protein synthesis and does not require evoked neurotransmission, indicating that a change in the efficacy of spontaneous quantal release events is sufficient to trigger the induction of synaptic homeostasis. Finally, both the rapid induction and the sustained expression of synaptic homeostasis are blocked by mutations that disrupt the pore-forming subunit of the presynaptic CaV2.1 calcium channel encoded by cacophony. These data confirm the presynaptic expression of synaptic homeostasis and implicate presynaptic CaV2.1 in a homeostatic retrograde signaling system

Homeostatic Signaling at Central and Peripheral Synapses

The magnitude and timing of neural activity precisely determines adaptive responses to stimuli. Neural activity is subject to extensive transformation between sensation and motor execution, and this transformation itself is subject to change due to the rewiring of the nervous system during development and learning. In this thesis I present data exploring how synapses between neural circuit elements might be homeostatically regulated, such that activity levels are maintained to ensure effective functioning of the nervous system. Inhibition of postsynaptic glutamate receptors at the Drosophila neuromuscular junction (NMJ) initiates a compensatory increase in presynaptic release termed synaptic homeostasis. This ensures that muscle depolarization stays constant despite diminished postsynaptic function, and this process of synaptic homeostasis may be important for maintaining proper NMJ function throughout development. While BMP signaling has been proposed to mediate the retrograde signal that controls synaptic homeostasis at this synapse, BMP signaling is also necessary for normal synaptic growth and stability. It remains unknown whether BMPs function as instructive retrograde signals that directly modulate presynaptic transmitter release. Here we demonstrate that the BMP receptor (Wit) and ligand (Gbb) are necessary for the induction of synaptic homeostasis, but that Gbb does not function as an instructive retrograde signal. Rather our data indicate that Wit and Gbb function via the downstream transcription factor Mad, and that Mad-mediated signaling is continuously required to gate the expression of synaptic homeostasis in motoneurons. Forms of synaptic plasticity in the central nervous system such as long-term potentiation (LTP) increase synaptic activity in a synapse- and cell-specific fashion. Although network-wide excitation triggers compensatory homeostatic changes, whether vertebrate neurons initiate homeostatic synaptic changes in response to cell-autonomous increases in excitation has not been examined. We cell-autonomously excited rodent CA1 pyramidal neurons and find that a compensatory postsynaptic depression of both AMPAR and NMDAR function results. Elevated calcium influx through L-type calcium channels leads to activation of a pathway involving CaM kinase kinase and CaM kinase 4 that induces synaptic depression of AMPAR and NMDAR responses. The synaptic depression of AMPARs but not of NMDARs requires protein synthesis and the GluA2 AMPAR subunit

2001. Regions in rat and human parathyroid hormone (PTH) 2 receptors controlling receptor interaction with PTH and with antagonist ligands. J Pharmacol Exp Ther

ABSTRACT The parathyroid hormone (PTH) 2 receptor is potently activated by tuberoinfundibular peptide (TIP39). Rat and human PTH2 receptors differ considerably in their PTH responsiveness. PTH weakly stimulates cAMP accumulation via the rat receptor, and here we show it did not detectably increase intracellular calcium ([Ca 2ϩ ] i ) and bound with low affinity (450 nM). For the human PTH2 receptor PTH was a full agonist for increasing cAMP, a partial agonist for increasing [Ca 2ϩ ] i , and bound with high affinity (18 nM). In addition, the antagonists PTH(7-34) and TIP(7-39) bound with 10-to 49-fold lower affinity to the rat receptor. We investigated the molecular basis of differential PTH and antagonist interaction with human and rat PTH2 receptors by using chimeric human/rat PTH2 receptors. PTH cAMP-signaling efficacy (E max ) was determined by extracellular loop (EL) 1 and a region including EL2 and EL3. The N-terminal domain determined PTH binding selectivity at the inactive receptor state. Multiple regions throughout the receptor are required for the PTH-PTH2 receptor complex to adopt a highaffinity active state: inserting the rat receptor's N-terminal domain, EL1 or EL2/3, into the human receptor increased PTH's EC 50 and reciprocal exchanges did not reduce EC 50 . This suggests the global receptor conformation prevents the rat receptor from adopting a high-affinity state when in complex with PTH. N-terminal ligand truncation, producing the antagonists PTH(7-34) and TIP(7-39), altered ligand interaction with the membrane-embedded domain of the receptor, eliminating EL2/3 as a specificity determinant and lowering binding affinity. These insights should contribute to the development of a highaffinity PTH2 receptor antagonist, for investigating the receptor's physiological role. The parathyroid hormone (PTH) 2 receptor from various species (human, rat, and zebrafish) is potently activated by a recently identified neuropeptide, tuberoinfundibular peptide of 39 residues (TIP39) The PTH2 receptor and TIP39 form part of an extended family of related receptors and related ligands that also includes the PTH1 receptor, PTH, and PTH-related protein (PTHrP) . The ligands and receptors have presumably evolved to selectively mediate different physiological functions. In this regard, the PTH2 receptor is potently activated by TIP39 but not by PTHrP ) and the PTH1 receptor is activated by PTH and PTHrP but not by TIP3

A deleterious Nav1.1 mutation selectively impairs telencephalic inhibitory neurons derived from Dravet Syndrome patients

Dravet Syndrome is an intractable form of childhood epilepsy associated with deleterious mutations in SCN1A, the gene encoding neuronal sodium channel Nav1.1. Earlier studies using human induced pluripotent stem cells (iPSCs) have produced mixed results regarding the importance of Nav1.1 in human inhibitory versus excitatory neurons. We studied a Nav1.1 mutation (p.S1328P) identified in a pair of twins with Dravet Syndrome and generated iPSC-derived neurons from these patients. Characterization of the mutant channel revealed a decrease in current amplitude and hypersensitivity to steady-state inactivation. We then differentiated Dravet-Syndrome and control iPSCs into telencephalic excitatory neurons or medial ganglionic eminence (MGE)-like inhibitory neurons. Dravet inhibitory neurons showed deficits in sodium currents and action potential firing, which were rescued by a Nav1.1 transgene, whereas Dravet excitatory neurons were normal. Our study identifies biophysical impairments underlying a deleterious Nav1.1 mutation and supports the hypothesis that Dravet Syndrome arises from defective inhibitory neurons

Behavioral abnormalities and circuit defects in the basal ganglia of a mouse model of 16p11.2 deletion syndrome.

A deletion on human chromosome 16p11.2 is associated with autism spectrum disorders. We deleted the syntenic region on mouse chromosome 7F3. MRI and high-throughput single-cell transcriptomics revealed anatomical and cellular abnormalities, particularly in cortex and striatum of juvenile mutant mice (16p11(+/-)). We found elevated numbers of striatal medium spiny neurons (MSNs) expressing the dopamine D2 receptor (Drd2(+)) and fewer dopamine-sensitive (Drd1(+)) neurons in deep layers of cortex. Electrophysiological recordings of Drd2(+) MSN revealed synaptic defects, suggesting abnormal basal ganglia circuitry function in 16p11(+/-) mice. This is further supported by behavioral experiments showing hyperactivity, circling, and deficits in movement control. Strikingly, 16p11(+/-) mice showed a complete lack of habituation reminiscent of what is observed in some autistic individuals. Our findings unveil a fundamental role of genes affected by the 16p11.2 deletion in establishing the basal ganglia circuitry and provide insights in the pathophysiology of autism

Behavioral Abnormalities and Circuit Defects in the Basal Ganglia of a Mouse Model of 16p11.2 Deletion Syndrome

A deletion on human chromosome 16p11.2 is associated with autism spectrum disorders. We deleted the syntenic region on mouse chromosome 7F3. MRI and high-throughput single-cell transcriptomics revealed anatomical and cellular abnormalities, particularly in cortex and striatum of juvenile mutant mice (16p11+/−). We found elevated numbers of striatal medium spiny neurons (MSNs) expressing the dopamine D2 receptor (Drd2+) and fewer dopamine-sensitive (Drd1+) neurons in deep layers of cortex. Electrophysiological recordings of Drd2+ MSN revealed synaptic defects, suggesting abnormal basal ganglia circuitry function in 16p11+/− mice. This is further supported by behavioral experiments showing hyperactivity, circling, and deficits in movement control. Strikingly, 16p11+/− mice showed a complete lack of habituation reminiscent of what is observed in some autistic individuals. Our findings unveil a fundamental role of genes affected by the 16p11.2 deletion in establishing the basal ganglia circuitry and provide insights in the pathophysiology of autism

Neuronal defects in a human cellular model of 22q11.2 deletion syndrome

22q11.2 deletion syndrome (22q11DS) is a highly penetrant and common genetic cause of neuropsychiatric disease. Here we generated induced pluripotent stem cells from 15 individuals with 22q11DS and 15 control individuals and differentiated them into three-dimensional (3D) cerebral cortical organoids. Transcriptional profiling across 100 days showed high reliability of differentiation and revealed changes in neuronal excitability-related genes. Using electrophysiology and live imaging, we identified defects in spontaneous neuronal activity and calcium signaling in both organoid- and 2D-derived cortical neurons. The calcium deficit was related to resting membrane potential changes that led to abnormal inactivation of voltage-gated calcium channels. Heterozygous loss of DGCR8 recapitulated the excitability and calcium phenotypes and its overexpression rescued these defects. Moreover, the 22q11DS calcium abnormality could also be restored by application of antipsychotics. Taken together, our study illustrates how stem cell derived models can be used to uncover and rescue cellular phenotypes associated with genetic forms of neuropsychiatric disease

Cerebrospinal fluid concentration of complement component 4A is increased in first episode schizophrenia.

Postsynaptic density is reduced in schizophrenia, and risk variants increasing complement component 4A (C4A) gene expression are linked to excessive synapse elimination. In two independent cohorts, we show that cerebrospinal fluid (CSF) C4A concentration is elevated in patients with first-episode psychosis (FEP) who develop schizophrenia (FEP-SCZ: median 0.41 fmol/ul [CI = 0.34-0.45], FEP-non-SCZ: median 0.29 fmol/ul [CI = 0.22-0.35], healthy controls: median 0.28 [CI = 0.24-0.33]). We show that the CSF elevation of C4A in FEP-SCZ exceeds what can be expected from genetic risk variance in the C4 locus, and in patient-derived cellular models we identify a mechanism dependent on the disease-associated cytokines interleukin (IL)-1beta and IL-6 to selectively increase neuronal C4A mRNA expression. In patient-derived CSF, we confirm that IL-1beta correlates with C4A controlled for genetically predicted C4A RNA expression (r = 0.39; CI: 0.01-0.68). These results suggest a role of C4A in early schizophrenia pathophysiology