Dissecting the mechanisms of synaptic transmission touches on nearly all fields of neuroscience. Work over recent years has made clear that protein distribution within single synapses is highly organized across multiple spatial scales ranging from the nanoscale accumulation of just a few protein molecules to larger domains with unique multiprotein compositions. I will address recent data regarding synaptic molecular organization to argue that the complexity of synaptic nanostructure generates functional capabilities that can fine-tune synaptic strength and augment the capabilities of classical quantal synaptic transmission. PSD-95 serves a case study for how to approach the emergent problem of describing and classifying forms of protein organization, including trans-synaptic “nanocolumn” relationships between AMPA receptors and glutamate release sites. Recent work identifying features of NMDA receptor subsynaptic organization suggests that these patterns also are likely to regulate the patterns of neural activity which can induce synaptic plasticity. The capacity of next-generation light microscopy to measure within synapses at sub-molecular resolution offers new routes to understanding the link between biochemical mechanisms and synapse physiology.