Intracellular recordings were made from neurones located in the ventromedial hypothalamic nucleus (VMHN) of slices from rat hypothalamus. These neurones were hyperpolarized on removal of extracellular glucose, resulting in an inhibition of firing, actions which were reversed on the re-introduction of glucose. No reversal of the inhibition of firing was observed when 10 mM mannoheptulose, an inhibitor of glucose metabolism, was present in addition to glucose. Increasing the mannoheptulose concentration to 20 mM resulted in further hyperpolarization. Cell-attached recordings from isolated neurones revealed that an increase in extracellular glucose inhibited a K+ channel and increased action current activity. ATP induced closure of this K+ channel when applied to inside-out membrane patches. Closure was also induced by Mg-free ATP or the non-hydrolysable ATP-analogue, adenylylimidodiphosphate indicating no requirement for ATP metabolism. We suggest that the closure of ATP-sensitive potassium channels underlies increased hypothalamic firing following an increase in extracellular glucose.

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1 The sulphonylureas, tolbutamide (0.1-10mM) and glibenclamide (0.1-100/pM) were shown not to inhibit ATP-K+ channel currents when applied to inside-out membrane patches excised from rat cultured cerebral cortex or freshly-dispersed ventromedial hypothalmic nucleus (VMHN) neurones. 2 Saturable binding sites for [3H]-glibenclamide, with similar affinity constants are present in rat cerebral cortex and hypothalamic membranes. The density of binding sites was lower in the hypothalamus than cortex. 3 Intracellular recordings from glucoreceptive VMHN neurones in hypothalamic slices were obtained. In the absence of glucose, tolbutamide (0.1 mM) depolarized these cells, increased membrane resistance and elicited action potentials. 4 Tolbutamide (0.1 mM) inhibited ATP-K+ channel currents and induced action current activity in cellattached recordings from glucoreceptive VMHN neurones. 5 Glibenclamide (10-500nM) had no effect per se on glucoreceptive VMHN neurones but did antagonize the actions of tolbutamide. 6 It is concluded that the hypothalamic (and perhaps cortical) sulphonylurea receptors are not directly coupled to ATP-K+ channels.

Oligomeric forms of β-amyloid (Aβ) have potent neurotoxic activity and are the primary cause of neuronal injury and cell death in Alzheimer's disease (AD). Compounds that perturb oligomer formation or structure may therefore be therapeutic for AD. We previously reported that d-[(chGly)-(Tyr)-(chGly)-(chGly)-(mLeu)]-NH(2) (SEN304) is able to inhibit Aβ aggregation and toxicity, shown primarily by thioflavin T fluorescence and MTT (Kokkoni, N. et al. (2006) N-Methylated peptide inhibitors of β-amyloid aggregation and toxicity. Optimisation of inhibitor structure. Biochemistry 45, 9906-9918). Here we extensively characterize how SEN304 affects Aβ(1-42) aggregation and toxicity, using biophysical assays (thioflavin T, circular dichroism, SDS-PAGE, size exclusion chromatography, surface plasmon resonance, traveling wave ion mobility mass spectrometry, electron microscopy, ELISA), toxicity assays in cell culture (MTT and lactate dehydrogenase in human SH-SHY5Y cells, mouse neuronal cell death and synaptophysin) and long-term potentiation in a rat hippocampal brain slice. These data, with dose response curves, show that SEN304 is a powerful inhibitor of Aβ(1-42) toxicity, particularly effective at preventing Aβ inhibition of long-term potentiation. It can bind directly to Aβ(1-42), delay β-sheet formation and promote aggregation of toxic oligomers into a nontoxic form, with a different morphology that cannot bind thioflavin T. SEN304 appears to work by inducing aggregation, and hence removal, of Aβ oligomers. It is therefore a promising lead compound for Alzheimer's disease.

The ability of neurons in the central nervous system to grow through a lesion and restore conduction has been analyzed in a developing spinal cord. The preparation consists of the entire central nervous system of the newly born opossum (Monodeiphis domestica), isolated and maintained in culture. Cell division, cell migration, and reflexes are maintained in such preparations for up to 8 days in culture. In the present experiments, massive lesions were produced by crushing the spinal cord, which abolished all conduction for a day. By 2-3 days after injury, electrical conduction across the crush could be observed. After 4-5 days, clear recovery had occurred: the amplitude of the conducted volley was comparable to that in acute preparations. In such preparations, the spinal cord had largely regained its normal appearance at the crush site. Axons stained by carbocyanine dyes or horseradish peroxidase had, by 4 days, grown in profusion through the lesion and several millimeters beyond it. These experiments demonstrate that neurons in the central nervous system of newly born mammals, unlike those in adults, can respond to injury by rapid and extensive outgrowth in the absence of peripheral nerve bridges or antibodies that neutralize inhibitory factors of myelin. With rapid and reliable regeneration occurring in vitro, it becomes practicable to assay the effects of molecules that promote or inhibit the restoration of functional connections.Damage to the central nervous system (CNS) of adult mammals is usually followed by minimal regrowth or repair (1). Axons can regenerate and make connections with their targets only after grafts of peripheral nerves have been supplied (2-4) or after inhibitory growth molecules have been neutralized (5-7). Oligodendrocytes in particular have been shown to produce molecules that actively prevent neurons from growing. By contrast, neurons within the CNS of a developing mammalian embryo are unmyelinated, grow profusely, and form synapses in the virtual absence of glial cells. The aim of the present experiments was to determine whether damaged immature mammalian CNS might be better able to repair itself after an injury. Such tests have not been made on embryos in the past, for technical reasons. Although axonal outgrowth can occur in neonatal hamsters following injury to pyramidal-tract fibers, the regenerating axons are not able to grow across the lesion (8).A suitable experimental preparation for studying regeneration after an injury is provided by the central nervous system of the neonatal opossum (Monodelphis domestica). This animal, being a marsupial, is extremely immature at birth. The newly born pup has only rudimentary eyes, ears, and hindlimbs and is unable to walk or to right itself. Its CNS is incomplete and corresponds to that of a 14-day rat embryo, having no cerebellum and only a rudimentary forebrain (9). Yet this neonatal animal is able to suck, to breathe, and to perform selected vital functions. Earlier experiments (10-12) have shown that the CNS can be removed in ...