No abstract

The effects of neuronal density on morphological and functional attributes of the evolving networks were studied in cultured dissociated hippocampal neurons. Plating at different densities affected connectivity among the neurons, such that sparse networks exhibited stronger synaptic connections between pairs of recorded neurons. This was associated with different patterns of spontaneous network activity with enhanced burst size but reduced burst frequency in the sparse cultures. Neuronal density also affected the morphology of the dendrites and spines of these neurons, such that sparse neurons had a simpler dendritic tree and fewer dendritic spines. Additionally, analysis of neurons transfected with PSD95 revealed that in sparse cultures the synapses are formed on the dendritic shaft, whereas in dense cultures the synapses are formed primarily on spine heads. These experiments provide important clues on the role of neuronal density in population activity and should yield new insights into the rules governing neuronal network connectivity.

A fundamental issue in understanding activity-dependent long-term plasticity of neuronal networks is the interplay between excitatory and inhibitory synaptic drives in the network. Using dual whole-cell recordings in cultured hippocampal neurons, we examined synaptic changes occurring as a result of a transient activation of NMDA receptors in the network. This enhanced transient activation led to a long-lasting increase in synchrony of spontaneous activity of neurons in the network. Simultaneous long-term potentiation of excitatory synaptic strength and a pronounced long-term depression of inhibitory synaptic currents (LTDi) were produced, which were independent of changes in postsynaptic potential and Ca 2ϩ concentrations. Surprisingly, miniature inhibitory synaptic currents were not changed by the conditioning, whereas both frequency and amplitudes of miniature EPSCs were enhanced. LTDi was mediated by activation of a presynaptic GABA B receptor, because it was blocked by saclofen and CGP55845 [(2S)-3-{[(15)-1-(3,4-dichlorophenyl)ethyl]amino-2-hydroxypropyl)(phenylmethyl)phosphinic acid].The cAMP antagonist Rp-adenosine 3Ј,5Ј-cyclic monophosphothioate abolished all measured effects of NMDA-dependent conditioning, whereas a nitric oxide synthase inhibitor was ineffective. Finally, network-induced plasticity was not occluded by a previous spike-timing-induced plasticity, indicating that the two types of plasticity may not share the same mechanism. These results demonstrate that network plasticity involves opposite affects on inhibitory and excitatory neurotransmission.

A common denominator for the induction of morphological and functional plasticity in cultured hippocampal neurons involves the activation of excitatory synapses. We now demonstrate massive morphological plasticity in mature cultured hippocampal neurons caused by a brief exposure to glutamate. This plasticity involves a slow, 70%-80% increase in spine cross-section area associated with a significant reduction in the width of dendrites. These changes are age dependent and expressed only in cells >18 d in vitro (DIV). Activation of both NMDARs and AMPARs as well as a sustained rise of internal calcium levels are necessary for induction of this plasticity. On the other hand, blockade of network activity or mGluRs does not abolish the observed morphological plasticity. Electrophysiologically, a brief exposure to glutamate induces an increase in the magnitude of EPSCs evoked between pairs of neurons, as well as in mEPSC frequency and amplitude, in mature but not young cultures. These results demonstrate an age-dependent, rapid and robust morphological and functional change in cultured central neurons that may contribute to their ability to express long term synaptic plasticity.