N -methyl- d -aspartate receptors (NMDARs) mediate critical CNS functions, whereas excessive activity contributes to neuronal damage. At physiological glycine concentrations, NMDAR currents recorded from cultured rodent hippocampal neurons exhibited strong desensitization in the continued presence of NMDA, thus protecting neurons from calcium overload. Reducing copper availability by specific chelators (bathocuproine disulfonate, cuprizone) induced nondesensitizing NMDAR currents even at physiologically low glycine concentrations. This effect was mimicked by, and was not additive with, genetic ablation of cellular prion protein (PrP C ), a key copper-binding protein in the CNS. Acute ablation of PrP C by enzymatically cleaving its cell-surface GPI anchor yielded similar effects. Biochemical studies and electrophysiological measurements revealed that PrP C interacts with the NMDAR complex in a copper-dependent manner to allosterically reduce glycine affinity for the receptor. Synthetic human Aβ 1–42 (10 nM–5 μM) produced an identical effect that could be mitigated by addition of excess copper ions or NMDAR blockers. Taken together, Aβ 1–42 , copper chelators, or PrP C inactivation all enhance the activity of glycine at the NMDAR, giving rise to pathologically large nondesensitizing steady-state NMDAR currents and neurotoxicity. We propose a physiological role for PrP C , one that limits excessive NMDAR activity that might otherwise promote neuronal damage. In addition, we provide a unifying molecular mechanism whereby toxic species of Aβ 1–42 might mediate neuronal and synaptic injury, at least in part, by disrupting the normal copper-mediated, PrP C -dependent inhibition of excessive activity of this highly calcium-permeable glutamate receptor.

It is well established that misfolded forms of cellular prion protein (PrP [PrPC]) are crucial in the genesis and progression of transmissible spongiform encephalitis, whereas the function of native PrPC remains incompletely understood. To determine the physiological role of PrPC, we examine the neurophysiological properties of hippocampal neurons isolated from PrP-null mice. We show that PrP-null mouse neurons exhibit enhanced and drastically prolonged N-methyl-d-aspartate (NMDA)–evoked currents as a result of a functional upregulation of NMDA receptors (NMDARs) containing NR2D subunits. These effects are phenocopied by RNA interference and are rescued upon the overexpression of exogenous PrPC. The enhanced NMDAR activity results in an increase in neuronal excitability as well as enhanced glutamate excitotoxicity both in vitro and in vivo. Thus, native PrPC mediates an important neuroprotective role by virtue of its ability to inhibit NR2D subunits.

Kv4 low voltage-activated A-type potassium channels are widely expressed in excitable cells, where they control action potential firing, dendritic activity and synaptic integration. Kv4 channels exist as a complex that includes K(+) channel-interacting proteins (KChIPs), which contain calcium-binding domains and therefore have the potential to confer calcium dependence on the Kv4 channel. We found that T-type calcium channels and Kv4 channels form a signaling complex in rat that efficiently couples calcium influx to KChIP3 to modulate Kv4 function. This interaction was critical for allowing Kv4 channels to function in the subthreshold membrane potential range to regulate neuronal firing properties. The widespread expression of these channels and accessory proteins indicates that the Cav3-Kv4 signaling complex is important for the function of a wide range of electrically excitable cells.

The inhibition of N-type calcium channels by opioid receptor like receptor 1 (ORL1) is a key mechanism for controlling the transmission of nociceptive signals. We recently reported that signaling complexes consisting of ORL1 receptors and N-type channels mediate a tonic inhibition of calcium entry. Here we show that prolonged ( approximately 30 min) exposure of ORL1 receptors to their agonist nociceptin triggers an internalization of these signaling complexes into vesicular compartments. This effect is dependent on protein kinase C activation, occurs selectively for N-type channels and cannot be observed with mu-opioid or angiotensin receptors. In expression systems and in rat dorsal root ganglion neurons, the nociceptin-mediated internalization of the channels is accompanied by a significant downregulation of calcium entry, which parallels the selective removal of N-type calcium channels from the plasma membrane. This may provide a new means for long-term regulation of calcium entry in the pain pathway.

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Control over the frequency and pattern of neuronal spike discharge depends on Ca2+-gated K+ channels that reduce cell excitability by hyperpolarizing the membrane potential. The Ca2+-dependent slow afterhyperpolarization (sAHP) is one of the most prominent inhibitory responses in the brain, with sAHP amplitude linked to a host of circuit and behavioral functions, yet the channel that underlies the sAHP has defied identification for decades. Here, we show that intermediate-conductance Ca2+-dependent K+ (IKCa) channels underlie the sAHP generated by trains of synaptic input or postsynaptic stimuli in CA1 hippocampal pyramidal cells. These findings are significant in providing a molecular identity for the sAHP of central neurons that will identify pharmacological tools capable of potentially modifying the several behavioral or disease states associated with the sAHP.

Dopamine signaling through D1 receptors in the prefrontal cortex (PFC) plays a critical role in the maintenance of higher cognitive functions, such as working memory. At the cellular level, these functions are predicated to involve alterations in neuronal calcium levels. The dendrites of PFC neurons express D1 receptors and N-type calcium channels, yet little information exists regarding their coupling. Here, we show that D1 receptors potently inhibit N-type channels in dendrites of rat PFC neurons. Using coimmunoprecipitation, we demonstrate the existence of a D1 receptor-N-type channel signaling complex in this region, and we provide evidence for a direct receptor-channel interaction. Finally, we demonstrate the importance of this complex to receptor-channel colocalization in heterologous systems and in PFC neurons. Our data indicate that the N-type calcium channel is an important physiological target of D1 receptors and reveal a mechanism for D1 receptor-mediated regulation of cognitive function in the PFC.

Encoding sensory input requires the expression of postsynaptic ion channels to transform key features of afferent input to an appropriate pattern of spike output. Although Ca 2+ -activated K + channels are known to control spike frequency in central neurons, Ca 2+ -activated K + channels of intermediate conductance (KCa3.1) are believed to be restricted to peripheral neurons. We now report that cerebellar Purkinje cells express KCa3.1 channels, as evidenced through single-cell RT-PCR, immunocytochemistry, pharmacology, and single-channel recordings. Furthermore, KCa3.1 channels coimmunoprecipitate and interact with low voltage-activated Cav3.2 Ca 2+ channels at the nanodomain level to support a previously undescribed transient voltage-and Ca 2+ -dependent current. As a result, subthreshold parallel fiber excitatory postsynaptic potentials (EPSPs) activate Cav3 Ca 2+ influx to trigger a KCa3.1-mediated regulation of the EPSP and subsequent after-hyperpolarization. The Cav3-KCa3.1 complex provides powerful control over temporal summation of EPSPs, effectively suppressing low frequencies of parallel fiber input. KCa3.1 channels thus contribute to a high-pass filter that allows Purkinje cells to respond preferentially to high-frequency parallel fiber bursts characteristic of sensory input.C entral neurons receive an enormous number of spontaneously active synaptic inputs, but exhibit the capacity to differentiate features of sensory input from background noise. Cerebellar Purkinje cells are contacted by up to ∼150,000 parallel fibers from granule cells, of which only a subset will convey sensory information at any given time. The activation of a peripheral receptive field is transmitted to the cerebellar cortex by mossy fibers in the form of high-frequency spike bursts (1). The resulting temporal summation of excitatory postsynaptic potentials (EPSPs) generates a similar high-frequency burst in granule cells (2). Purkinje cells should then also possess the means to respond effectively to bursts of parallel fiber input that convey sensory information compared with background activity.Postsynaptic membrane excitability can be controlled by activation of K + channels. There are two established types of Ca 2+ -activated K + (KCa) channels in CNS neurons: small conductance (SK, KCa2.x) and big conductance (BK, KCa1.1) (3, 4). A third class of intermediate conductance (KCa3.1, SK4, IK1) KCa channel is thought to be expressed only in microglia and endothelial cells in the CNS (3, 5, 6). KCa3.1 channels are gated by calmodulin in a similar manner to KCa2.x channels but are insensitive to block by apamin and tetraethylammonium (TEA) (6-8). Instead, the KCa3.1 α-subunit, encoded by the gene KCNN4, has specific residues that bind charybdotoxin and 1-[(2-chlorophenyl) diphenylmethyl]-1H-pyrazole (TRAM-34) (5-7, 9, 10).In cerebellar Purkinje cells, KCa1.1 and KCa2.2 channels are activated during a spike by high voltage-activated (HVA) P-type Ca 2+ channels (11). In contrast, low voltage-activated (LVA) Cav3 (T-type) Ca 2+ channe...