Carlos D. Aizenman

Rapid, synaptically driven increases in the intrinsic excitability ofcerebellar deep nuclear neurons

The neurons of the cerebellar deep nuclei are implicated in certain forms of motor learning such as associative eyeblink conditioning, partly because increases in their firing rates parallel acquisition of the conditioned response. Here we demonstrate that these neurons can show persistent increases in their intrinsic excitability following a Ca2+ load imposed by synaptic activation of NMDA receptors or direct current injection. This phenomenon, together with use-dependent alterations in synaptic strength, may provide a flexible and informationally rich engram for cerebellar motor learning.

Polarity of Long-Term Synaptic Gain Change Is Related to Postsynaptic Spike Firing at a Cerebellar Inhibitory Synapse

Long-term potentiation and depression (LTP and LTD) in excitatory synapses can coexist, the former being triggered by stimuli that produce strong postsynaptic excitation and the latter by stimuli that produce weaker postsynaptic excitation. It has not been determined whether these properties also apply to LTP and LTD in the inhibitory synapses between Purkinje neurons and the neurons of the deep cerebellar nuclei (DCN), a site that has been implicated in certain types of motor learning. DCN cells exhibit a prominent rebound depolarization (RD) and associated spike burst upon release from hyperpolarization. In these cells, LTP can be elicited by short, high-frequency trains of inhibitory postsynaptic potentials (IPSPs), which reliably evoke an RD. LTD is induced if the same protocol is applied with conditions where the amount of postsynaptic excitation is reduced. The polarity of the change in synaptic strength is correlated with the amount of RD-evoked spike firing during the induction protocol. Thus, some important computational principles that govern the induction of use-dependent change in excitatory synaptic efficacy also apply to inhibitory synapses.

Regulation of rho GTPases by crosstalk and neuronal activity in vivo.

Proper development of neurons depends on synaptic activity, but the mechanisms of activity-dependent neuronal growth are not well understood. The small GTPases, RhoA, Rac, and Cdc42, regulate neuronal morphogenesis by controlling the assembly and stability of the actin cytoskeleton. We report an in situ method to determine endogenous Rho GTPase activity in intact Xenopus brain. We use this method to provide evidence for crosstalk between Rho GTPases in optic tectal cells. Moreover, crosstalk between the Rho GTPases appears to affect dendritic arbor development in vivo. Finally, we demonstrate that optic nerve stimulation regulates Rho GTPase activity in a glutamate receptor-dependent manner. These data suggest a link between glutamate receptor function, Rho GTPase activity, and dendritic arbor growth in the intact animal.

Visually driven regulation of intrinsic neuronal excitability improves stimulus detection in vivo.

Neurons adapt their electrophysiological properties to maintain stable levels of electrical excitability when faced with a constantly changing environment. We find that exposing freely swimming Xenopus tadpoles to 4-5 hr of persistent visual stimulation increases the intrinsic excitability of optic tectal neurons. This increase is correlated with enhanced voltage-gated Na+ currents. The same visual stimulation protocol also induces a polyamine synthesis-dependent reduction in Ca2+-permeable AMPAR-mediated synaptic drive, suggesting that the increased excitability may compensate for this reduction. Accordingly, the change in excitability was prevented by blocking polyamine synthesis during visual stimulation and was rescued when Ca2+-permeable AMPAR-mediated transmission was selectively reduced. The changes in excitability also rendered tectal cells more responsive to synaptic burst stimuli, improving visual stimulus detection. The synaptic and intrinsic adaptations function together to keep tectal neurons within a constant operating range, while making the intact visual system less responsive to background activity yet more sensitive to burst stimuli.

Homeostatic Regulation of Intrinsic Excitability and Synaptic Transmission in a Developing Visual Circuit

One of the major challenges faced by the developing visual system is how to stably process visual information, yet at the same time remain flexible enough to accommodate growth and plasticity induced by visual experience. We find that in the Xenopus retinotectal circuit, during a period in development when the retinotectal map undergoes activity-dependent refinement and visual inputs strengthen, tectal neurons adapt their intrinsic excitability such that a stable relationship between the total level of synaptic input and tectal neuron spike output is conserved. This homeostatic balance between synaptic and intrinsic properties is maintained, in part, via regulation of voltage-gated Na+ currents, resulting in a stable neuronal input–output function. We experimentally manipulated intrinsic excitability or synapse strengthening in developing tectal neurons in vivo by electroporation of a leak K+ channel gene or a peptide that interferes with normal AMPA receptor trafficking. Both manipulations resulted in a compensatory increase in voltage-gated Na+ currents. This suggests that intrinsic neuronal properties are actively regulated as a function of the total level of neuronal activity experienced during development. We conclude that the coordinated changes between synaptic and intrinsic properties allow developing optic tectal neurons to remain within a stable dynamic range, even as the pattern and strength of visual inputs changes over development, suggesting that homeostatic regulation of intrinsic properties plays a central role in the functional development of neural circuits.

Visually Driven Modulation of Glutamatergic Synaptic Transmission Is Mediated by the Regulation of Intracellular Polyamines

Ca2+-permeable AMPARs are inwardly rectifying due to block by intracellular polyamines. Neuronal activity regulates polyamine synthesis, yet whether this affects Ca2+-AMPAR-mediated synaptic transmission is unknown. We test whether 4 hr of increased visual stimulation regulates glutamatergic retino-tectal synapses in Xenopus tadpoles. Tectal neurons containing Ca2+-AMPARs form a gradient along the rostro-caudal developmental axis. These neurons had inwardly rectifying AMPAR-mediated EPSCs. Four hours of visual stimulation or addition of intracellular spermine increased rectification in immature neurons. Polyamine synthesis inhibitors blocked the effect of visual stimulation, suggesting that visual activity regulates AMPARs via the polyamine synthesis pathway. This modulation resulted in changes in the integrative properties of tectal neurons. Regulation of polyamine synthesis by physiological stimuli is a novel form of modulation of synaptic transmission important for understanding the short-term effects of enhanced sensory experience during development.

Roles of NR2A and NR2B in the Development of Dendritic Arbor Morphology In Vivo

NMDA receptors (NMDARs) are important for neuronal development and circuit formation. The NMDAR subunits NR2A and NR2B are biophysically distinct and differentially expressed during development but their individual contribution to structural plasticity is unknown. Here we test whether NR2A and NR2B subunits have specific functions in the morphological development of tectal neurons in living Xenopus tadpoles. We use exogenous subunit expression and endogenous subunit knockdown to shift synaptic NMDAR composition toward NR2A or NR2B, as shown electrophysiologically. We analyzed the dendritic arbor structure and found evidence for both overlapping and distinct functions of NR2A and NR2B in dendritic development. Control neurons develop regions of high local branch density in their dendritic arbor, which may be important for processing topographically organized inputs. Exogenous expression of either NR2A or NR2B decreases local branch clusters, indicating a requirement for both subunits in dendritic arbor development. Knockdown of endogenous NR2A reduces local branch clusters, whereas knockdown of NR2B has no effect on branch clustering. Analysis of the underlying branch dynamics shows that exogenous NR2B-expressing neurons are more dynamic than control or exogenous NR2A-expressing neurons, demonstrating subunit-specific regulation of branch dynamics. Visual experience-dependent increases in dendritic arbor growth rate seen in control neurons are blocked in both exogenous NR2A- and NR2B-expressing neurons. These experiments indicate that NR2A and NR2B have subunit-specific properties in dendritic arbor development, but also overlapping functions, indicating a requirement for both subunits in neuronal development.

Development and spike timing-dependent plasticity of recurrent excitation in the Xenopus optic tectum.

Much of the information processing in the brain occurs at the level of local circuits; however, the mechanisms underlying their initial development are poorly understood. We sought to examine the early development and plasticity of local excitatory circuits in the optic tectum of Xenopus laevis tadpoles. We found that retinal input recruits persistent, recurrent intratectal synaptic excitation that becomes more temporally compact and less variable over development, thus increasing the temporal coherence and precision of tectal cell spiking. We also saw that patterned retinal input can sculpt recurrent activity according to a spike timing–dependent plasticity rule, and that impairing this plasticity during development results in abnormal refinement of the temporal characteristics of recurrent circuits. This plasticity is a previously unknown mechanism by which patterned retinal activity allows intratectal circuitry to self-organize, optimizing the temporal response properties of the tectal network, and provides a substrate for rapid modulation of tectal neuron receptive-field properties.

Visual Avoidance in Xenopus Tadpoles Is Correlated With the Maturation of Visual Responses in the Optic Tectum

The optic tectum is central for transforming incoming visual input into orienting behavior. Yet it is not well understood how this behavior is organized early in development and how it relates to the response properties of the developing visual system. We designed a novel behavioral assay to study the development of visually guided behavior in Xenopus laevis tadpoles. We found that, during early development, visual avoidance-an innate, tectally mediated behavior-is tuned to a specific stimulus size and is sensitive to changes in contrast. Using in vivo recordings we found that developmental changes in the spatial tuning of visual avoidance are mirrored by changes in tectal receptive field sharpness and the temporal properties of subthreshold visual responses, whereas contrast sensitivity is affected by the gain of the visual response. We also show that long- and short-term perturbations of visual response properties predictably alter behavioral output. We conclude that our assay for visual avoidance is a useful functional measure of the developmental state of the tectal circuitry. We use this assay to show that the developing visual system is tuned to facilitate behavioral output and that the system can be modulated by neural activity, allowing it to adapt to environmental changes it encounters during development.

Neurodevelopmental effects of chronic exposure to elevated levels of pro-inflammatory cytokines in a developing visual system

BackgroundImbalances in the regulation of pro-inflammatory cytokines have been increasingly correlated with a number of severe and prevalent neurodevelopmental disorders, including autism spectrum disorder, schizophrenia and Down syndrome. Although several studies have shown that cytokines have potent effects on neural function, their role in neural development is still poorly understood. In this study, we investigated the link between abnormal cytokine levels and neural development using the Xenopus laevis tadpole visual system, a model frequently used to examine the anatomical and functional development of neural circuits.ResultsUsing a test for a visually guided behavior that requires normal visual system development, we examined the long-term effects of prolonged developmental exposure to three pro-inflammatory cytokines with known neural functions: interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α. We found that all cytokines affected the development of normal visually guided behavior. Neuroanatomical imaging of the visual projection showed that none of the cytokines caused any gross abnormalities in the anatomical organization of this projection, suggesting that they may be acting at the level of neuronal microcircuits. We further tested the effects of TNF-α on the electrophysiological properties of the retinotectal circuit and found that long-term developmental exposure to TNF-α resulted in enhanced spontaneous excitatory synaptic transmission in tectal neurons, increased AMPA/NMDA ratios of retinotectal synapses, and a decrease in the number of immature synapses containing only NMDA receptors, consistent with premature maturation and stabilization of these synapses. Local interconnectivity within the tectum also appeared to remain widespread, as shown by increased recurrent polysynaptic activity, and was similar to what is seen in more immature, less refined tectal circuits. TNF-α treatment also enhanced the overall growth of tectal cell dendrites. Finally, we found that TNF-α-reared tadpoles had increased susceptibility to pentylenetetrazol-induced seizures.ConclusionsTaken together our data are consistent with a model in which TNF-α causes premature stabilization of developing synapses within the tectum, therefore preventing normal refinement and synapse elimination that occurs during development, leading to increased local connectivity and epilepsy. This experimental model also provides an integrative approach to understanding the effects of cytokines on the development of neural circuits and may provide novel insights into the etiology underlying some neurodevelopmental disorders.