Abstracts

Rationale: Hypothalamic hamartomas (HH) are intrinsically epileptogenic for gelastic seizures, although the basic cellular mechanisms remain unknown. HH tissue consists of clusters of small interneuron-like neurons, and smaller numbers of large projection-type neurons, each with specific electrophysiological features, which together suggest a model for increased neuronal excitation. We have hypothesized that there is also increased synchrony within small neuron clusters, and that this synchrony results from abnormal non-synaptic cellular interactions. We predicted that evidence for abnormal non-synaptic connectivity would be apparent with thin-section electron microscopy. Here, we have examined the ultrastructure of subsurface cisterns (SSC), which occur immediately beneath the plasma membrane of neuron cell bodies. Normally, these micro-regions of the plasma membrane possess differences in ionic conductance and channel expression relative to other regions of the soma membrane, and almost universally have intimate contact with adjacent astrocytic processes. SSCs also appear to have a role in intracellular signaling pathways as stores of intracellular calcium. Abnormalities of SSC have been linked to experimental epilepsy models. Methods: Surgically-resected HH tissue was utilized following informed consent under approved protocols. Thin (0.08um) sections from six patients were examined with a Philips CM 100 EM fitted with a digital camera. Single sections and 3-D reconstruction of serial sections of small HH neurons were used for analysis. Results: Most small neurons exhibited one or more SSCs, confined to the soma and proximal dendrites. HH SSCs appeared as discs composed of two collapsed or fused membranes of endoplasmic reticulum (ER) cisterns. The lumen of the cistern was essentially obliterated except at their lateral ends where a small canal remained. SSCs were 0.5 to 1.5 um in length, and always ran parallel with the neuronal plasma membrane. Dense granular material 15-30nm in thickness always connected SSCs to the plasma membrane. The neuronal plasma membrane domains overlying SSCs were significantly more electron dense compared to adjacent regions. Reconstruction of serial sections showed that only two of 24 SSCs were in close proximity to symmetrical (putative inhibitory) synapses. Abnormalities included absent or reduced coverage of the SSC-associated plasma membrane by astrocytic processes. A wide diversity of SSC/glia morphology was evident. Conclusions: SSCs of small HH neurons show abnormal ultrastructural features, with a lack of the expected relationship between the neuron and surrounding glia. SSCs are important regional sub-domains of membrane ionic conductance and sites of intracellular signaling. Abnormalities of SSC function may contribute to altered excitability or synchrony of HH tissue. This is the first study to implicate glial mechanisms as a possible component of seizure genesis within HH tissue.