Children who survive preterm birth exhibit persistent unexplained disturbances in cerebral cortical growth with associated cognitive and learning disabilities. The mechanisms underlying these deficits remain elusive. We used ex vivo diffusion magnetic resonance imaging to demonstrate in a preterm large-animal model that cerebral ischemia impairs cortical growth and the normal maturational decline in cortical fractional anisotropy (FA). Analysis of pyramidal neurons revealed that cortical deficits were associated with impaired expansion of the dendritic arbor and reduced synaptic density. Together, these findings suggest a link between abnormal cortical FA and disturbances of neuronal morphological development. To experimentally investigate this possibility, we measured the orientation distribution of dendritic branches and observed that it corresponds with the theoretically predicted pattern of increased anisotropy within cases that exhibited elevated cortical FA after ischemia. We conclude that cortical growth impairments are associated with diffuse disturbances in the dendritic arbor and synapse formation of cortical neurons, which may underlie the cognitive and learning disabilities in survivors of preterm birth. Further, measurement of cortical FA may be useful for noninvasively detecting neurological disorders affecting cortical development.

Delta-conotoxins are Conus peptides that inhibit inactivation of voltage-gated sodium channels. The suggestion that delta-conotoxins might be an essential component of the venoms of fish-hunting cone snails which rapidly immobilize their prey [Terlau, H., Shon, K., Grilley, M., Stocker, M., Stühmer, W., and Olivera, B. M. (1996) Nature 381, 148-151] has not been tested. On the basis of cDNA cloning, all of the fish-hunting Conus analyzed yielded at least one delta-conotoxin sequence. In addition, one delta-conotoxin isolated from the venom of Conus striatus had an amino acid sequence identical to that predicted from cDNA cloning. This new peptide exhibited properties of delta-conotoxins: it targeted sodium channels and potentiated action potentials by slowing channel inactivation. Homologous sequences of delta-conotoxins from two groups (clades) of related fish-hunting Conus species share consensus features but differ significantly from the two known delta-conotoxins from mollusc-hunting Conus venoms. Three large hydrophobic amino acids were conserved; analogues of the previously described delta-conotoxin PVIA with alanine substituted for the conserved amino acids F9 and I12 lost substantial biological activity. In contrast, both the T8A and K13A delta-conotoxin PVIA analogues, where substitutions were at nonconserved loci, proved to be biologically active. Taken together, our results indicate that a cladistic approach can identify amino acids critical for the activity of conotoxins and provide extensive information as to which amino acid substitutions can be made without significant functional consequences.

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