Cortical spheroids display oscillatory network dynamics.

Three-dimensional brain cultures can facilitate the study of central nervous system function and disease, and one of the most important components that they present is neuronal activity on a network level. Here we demonstrate network activity in rodent cortical spheroids while maintaining the networks intact in their 3D state. Networks developed by nine days in culture and became more complex over time. To measure network activity, we imaged neurons in rat and mouse spheroids labelled with a calcium indicator dye, and in mouse spheroids expressing GCaMP. Network activity was evident when we electrically stimulated spheroids, was abolished with glutamatergic blockade, and was altered by GABAergic blockade or partial glutamatergic blockade. We quantified correlations and distances between somas with micron-scale spatial resolution. Spheroids seeded at as few as 4000 cells gave rise to emergent network events, including oscillations. These results are the first demonstration that self-assembled rat and mouse spheroids exhibit network activity consistent with in vivo network events. These results open the door to experiments on neuronal networks that require fewer animals and enable high throughput experiments on network-perturbing alterations in neurons and glia.

A calcium-dependent pathway underlies activity-dependent plasticity of electrical synapses in the thalamic reticular nucleus.

KEY POINTS: Electrical synapses are modified by various forms of activity, including paired activity in coupled neurons and tetanization of the input to coupled neurons. We show that plasticity of electrical synapses that results from paired spiking activity in coupled neurons depends on calcium influx and calcium-initiated signalling pathways. Plasticity that results from tetanization of input fibres does not depend on calcium influx or dynamics. These results imply that electrically coupled neurons have distinct sets of mechanisms for adjusting coupling according to the specific type of activity they experience. ABSTRACT: Recent results have demonstrated modification of electrical synapse strength by varied forms of neuronal activity. However, the mechanisms underlying plasticity induction in central mammalian neurons are unclear. Here we show that the two established inductors of plasticity at electrical synapses in the thalamic reticular nucleus - paired burst spiking in coupled neurons, and mGluR-dependent tetanization of synaptic input - are separate pathways that converge at a common downstream endpoint. Using occlusion experiments and pharmacology in patched pairs of coupled neurons in vitro, we show that burst-induced depression depends on calcium entry via voltage-gated channels, is blocked by BAPTA chelation, and recruits intracellular calcium release on its way to activation of phosphatase activity. In contrast, mGluR-dependent plasticity is independent of calcium entry or calcium dynamics. Together, these results show that the spiking-initiated mechanisms underlying electrical synapse plasticity are similar to those that induce plasticity at chemical synapses, and offer the possibility that calcium-regulated mechanisms may also lead to alternate outcomes, such as potentiation. Because these mechanistic elements are widely found in mature neurons, we expect them to apply broadly to electrical synapses across the brain, acting as the crucial link between neuronal activity and electrical synapse strength.

Asymmetry and modulation of spike timing in electrically coupled neurons.

Electrical coupling mediates interactions between neurons of the thalamic reticular nucleus (TRN), which play a critical role in regulating thalamocortical and corticothalamic communication by inhibiting thalamic relay cells. Accumulating evidence has shown that asymmetry of electrical synapses is a fundamental and dynamic property, but the effect of asymmetry on coupled networks is unexplored. Recording from patched pairs in rat brain slices, we investigate asymmetry in the subthreshold regime and show that electrical synapses can exert powerful effects on the spike times of coupled neighbors. Electrical synaptic signaling modulates spike timing by 10-20 ms, in an effect that also exhibits asymmetry. Furthermore, we show through modeling that coupling asymmetry expands the set of outputs for pairs of coupled neurons through enhanced regions of synchrony and reversals of spike order. These results highlight the power and specificity of signaling exerted by electrical synapses, which contribute to information flow across the brain.