Guanine derivatives (GD) have been implicated in many relevant brain extracellular roles, such as modulation of glutamate transmission and neuronal protection against excitotoxic damage. GD are spontaneously released to the extracellular space from cultured astrocytes and during oxygen/glucose deprivation (OGD). The aim of this study has been to evaluate the potassium channels and phosphatidilinositol-3 kinase (PI3K) pathway involvement in the mechanisms related to the neuroprotective role of guanosine in rat hippocampal slices subjected to OGD. The addition of guanosine (100 μM) to hippocampal slices subjected to 15 min of OGD and followed by 2 h of re-oxygenation is neuroprotective. The presence of K+ channel blockers, glibenclamide (20 μM) or apamin (300 nM), revealed that neuroprotective effect of guanosine was not dependent on ATP-sensitive K+ channels or small conductance Ca²+-activated K+ channels. The presence of charybdotoxin (100 nM), a large conductance Ca²+-activated K+ channel (BK) blocker, inhibited the neuroprotective effect of guanosine. Hippocampal slices subjected to OGD and re-oxygenation showed a significant reduction of glutamate uptake. Addition of guanosine in the re-oxygenation period has blocked the reduction of glutamate uptake. This guanosine effect was inhibited when hippocampal slices were pre-incubated with charybdotoxin or wortmanin (a PI3K inhibitor, 1 μM) in the re-oxygenation period. Guanosine promoted an increase in Akt protein phosphorylation. However, the presence of charybdotoxin blocked such effect. In conclusion, the neuroprotective effect of guanosine involves augmentation of glutamate uptake, which is modulated by BK channels and the activation of PI3K pathway. Moreover, neuroprotection caused by guanosine depends on the increased expression of phospho-Akt protein.

DiGeorge/22q11.2 Deletion Syndrome (22q11DS) is a common genetic microdeletion syndrome that underlies several neurodevelopmental disorders including autism, attention deficit/hyperactivity disorder, and schizophrenia. In addition to cognitive impairments, those with 22q11DS have disrupted feeding and swallowing from birth onward. This perinatal dysphagia significantly compromises nutritional status, impairs appropriate weight gain, and can lead to life threatening aspiration-based infections. Appropriately timed excitation and inhibition of brainstem hypoglossal motor neurons, which innervate tongue muscles, is essential for proper feeding and swallowing. In this study we have examined changes in hypoglossal motor neuron function in the LgDel mouse model of 22q11DS. Hypoglossal motor neurons from LgDel mouse pups have action potentials with afterhyperpolarizations, mediated by a large conductance charybdotoxin-sensitive Ca-activated K current, that are significantly shorter in duration and greater in magnitude than those in wild-type pups. In addition, the amplitude, but not frequency, of glutamatergic excitatory glutamatergic postsynaptic currents (EPSCs) is diminished, and GABAergic, but not glycinergic, neurotransmission to hypoglossal motor neurons was reduced in LgDel animals. These observations provide a foundation for understanding the neurological changes in hypoglossal motor neuron function and their contribution to swallowing abnormalities that occur in DiGeorge/22q11.2 Deletion Syndrome.

Using Xenopus nerve-muscle co-cultures, we have examined the contribution of calcium-activated potassium (K(Ca)) channels to the regulation of transmitter release evoked by single action potentials. The presynaptic varicosities that form on muscle cells in these cultures were studied directly using patch-clamp recording techniques. In these developing synapses, blockade of K(Ca) channels with iberiotoxin or charybdotoxin decreased transmitter release by an average of 35%. This effect would be expected to be caused by changes in the late phases of action potential repolarization. We hypothesize that these changes are due to a reduction in the driving force for calcium that is normally enhanced by the local hyperpolarization at the active zone caused by potassium current through the K(Ca) channels that co-localize with calcium channels. In support of this hypothesis, we have shown that when action potential waveforms were used as voltage-clamp commands to elicit calcium current in varicosities, peak calcium current was reduced only when these waveforms were broadened beginning when action potential repolarization was 20% complete. In contrast to peak calcium current, total calcium influx was consistently increased following action potential broadening. A model, based on previously reported properties of ion channels, faithfully reproduced predicted effects on action potential repolarization and calcium currents. From these data, we suggest that the large-conductance K(Ca) channels expressed at presynaptic varicosities regulate transmitter release magnitude during single action potentials by altering the rate of action potential repolarization, and thus the magnitude of peak calcium current.