Coassembly of Big Conductance Ca2+-activated K+ Channels and L-type Voltage-gated Ca2+ Channels in Rat Brain

Abstract

Based on electrophysiological studies, Ca 2؉ -activated K ؉ channels and voltage-gated Ca 2؉ channels appear to be located in close proximity in neurons. Such colocalization would ensure selective and rapid activation of K ؉ channels by local increases in the cytosolic calcium concentration. The nature of the apparent coupling is not known. In the present study we report a direct coassembly of big conductance Ca 2؉ -activated K ؉ channels (BK) and L-type voltage-gated Ca 2؉ channels in rat brain. Saturation immunoprecipitation studies were performed on membranes labeled for BK channels and precipitated with antibodies against ␣ 1C and ␣ 1D L-type Ca 2؉ channels. To confirm the specificity of the interaction, precipitation experiments were carried out also in reverse order. Also, additive precipitation was performed because ␣ 1C and ␣ 1D L-type Ca 2؉ channels always refer to separate ion channel complexes. Finally, immunochemical studies showed a distinct but overlapping expression pattern of the two types of ion channels investigated. BK and L-type Ca 2؉ channels were colocalized in various compartments throughout the rat brain. Taken together, these results demonstrate a direct coassembly of BK channels and L-type Ca 2؉ channels in certain areas of the brain.

Co-assembly of N-type Ca2+ and BK channels underlies functional coupling in rat brain

“…1Ai) showed that Ca V 1.2 immunoreactivity was 45% weaker than Ca V 2.2 immunoreactivity. This weak interaction between L-type Ca 2+ and BK channels was reminiscent of the co-IP data reported by Grunnet and Kaufmann (Grunnet and Kaufmann, 2004). Thus, the data clearly indicated that the two channel subunits could associate in rat brain, but the association was more prevalent between Ntype Ca 2+ and BK channels.…”

“…Using reciprocal co-immunoprecipitation, we found that N-type channels were more abundantly associated with BK channels than L-type channels (Ca V 1.2) in rat brain. Expression of only the pore-forming ␣-subunits of the N-type (Ca V 2.2) and BK (Slo 27 ) channels in a non-neuronal cell-line gave robust macroscopic currents using biochemical techniques to demonstrate the association of L-type and BK channels in rat whole brain (Grunnet and Kaufmann, 2004;Berkefeld et al, 2006). However, no study has yet identified the rapid functional coupling between BK and Ca 2+ channels and resolved how this might be achieved.…”

“…2+ channels and BK channels Ca 2+ entry through both L-and N-type Ca 2+ channels activates BK channels in neocortical neurons (Sun et al, 2003) and association between L-type Ca 2+ and BK channels has been observed in rat whole brain (Grunnet and Kaufmann, 2004;Berkefeld et al, 2006). Immunoblotting an anti-BK␣ 1118-1135 co-IP (BK-IP) sample from rat brain with an antibody directed against the L-type Ca 2+ channel (anti-Ca V 1.2) revealed an immunoreactive band of the predicted molecular mass of the native Ca V 1.2 subunit (210 kDa).…”

Profiling the Phospho-status of the BKCa Channel α Subunit in Rat Brain Reveals Unexpected Patterns and Complexity

“…The C-DEC splice variant generates an extended C terminus resulting in BK␣ with enhanced retention of newly synthesized BK Ca channels in the endoplasmic reticulum, which finally results in decreased cell surface expression (60,61). BK Ca channels are robustly expressed in mammalian central neurons (62,63) and are predominantly localized to axons and presynaptic terminals (17,37) but are also present in certain neuronal somata and dendrites (18,64). In rat brain, BK␣ splice variants may be differentially expressed in distinct cell types or different compartments of the same neuron.…”

Prolactin Regulates Tuberoinfundibular Dopamine Neuron Discharge Pattern: Novel Feedback Control Mechanisms in the Lactotrophic Axis

“…These data suggest that Ca 2ϩ influx via L-type VDCCs is a requirement for the potassium current inhibited by Prl in the HV range. In neurons, BK channels often colocalize to microdomains with L-type Ca 2ϩ channels (Grunnet and Kaufmann, 2004;Berkefeld et al, 2006;Fakler and Adelman, 2008), and Ca 2ϩ influx through the latter is believed to shift BK voltage dependence to more hyperpolarized potentials. The tight coupling of L-type VDCCs to BK channels has been implicated in other endocrine and neuroendocrine cells with sustained secretion in response to steady depolarization (Prakriya and Lingle, 1999;Marcantoni et al, 2007Marcantoni et al, , 2010 and, in adrenomedullary cells, can be recruited as demands on hormone release increase (Polo-Parada et al, 2006).…”