The columnar organization is most apparent in the whisker barrel cortex but seems less apparent in the gustatory insular cortex. We addressed here whether there are any differences between the two cortices in columnar information processing by comparing the spatiotemporal patterns of excitation spread in the two cortices using voltage-sensitive dye imaging. In contrast to the well known excitation spread in the horizontal direction in layer II/III induced in the barrel cortex by layer IV stimulation, the excitation caused in the insular cortex by stimulation of layer IV spread bidirectionally in the vertical direction into layers II/III and V/VI, displaying a columnar image pattern. Bicuculline or picrotoxin markedly extended the horizontal excitation spread in layer II/III in the barrel cortex, leading to a generation of excitation in the underlying layer V/VI, whereas those markedly increased the amplitude of optical responses throughout the whole column in the insular cortex, subsequently widening the columnar image pattern. Such synchronous activities as revealed by the horizontal and vertical excitation spreads were consistently induced in the barrel and insular cortices, respectively, even by stimulation of different layers with varying intensities. Thus, a unique functional column existed in the insular cortex, in which intracolumnar communication between the superficial and deep layers was prominent, and GABA_Aaction is involved in the inhibition of the intracolumnar communication in contrast to its involvement in intercolumnar lateral inhibition in the barrel cortex. These results suggest that the columnar information processing may not be universal across the different cortical areas.

Noradrenergic neurons in the locus coeruleus (LC) release noradrenaline (NA) that acts via volume transmission to activate extrasynaptic G-protein coupled receptors (GPCRs) in target cells throughout the brain. As the closest projection, the dorsal LC laterally adjoins the mesencephalic trigeminal nucleus (MTN), in which proprioceptive primary sensory neurons innervating muscle spindles of jaw-closing muscles are exceptionally located. MTN neurons express α₂-adrenergic receptors (α₂-ARs) and display hyperpolarization-activated cyclic nucleotide-gated (HCN) currents (Ihs), which is downregulated by α₂-AR activation. To quantify the activity-dependent outcome of volume transmission of NA from LC to MTN, we investigated how direct LC activation inhibits Ih in MTN neurons by performing dual whole-cell recordings from LC and MTN neurons. Repetition of 20 Hz spike-train evoked with 1-s current-pulse in LC neurons every 30 s resulted in a gradual decrease in Ih evoked every 30 s, revealing a Hill-type relationship between the number of spike-trains in LC neurons and the degree of Ih inhibition in MTN neurons. On the other hand, when microstimulation was applied in LC every 30 s, an LC neuron repeatedly displayed a transient higher-frequency firing followed by a tonic firing at 5–10 Hz for 30 s. This subsequently caused a similar Hill-type inhibition of Ih in the simultaneously recorded MTN neuron, but with a smaller Hill coefficient, suggesting a lower signal transduction efficacy. In contrast, 20 Hz activity induced by a 1-s pulse applied every 5–10 s caused only a transient facilitation of Ih inhibition followed by a forced termination of Ih inhibition. Thus, the three modes of LC activities modulated the volume transmission to activate α₂-adrenergic GPCR to differentially inhibit Ih in MTN neurons.

The axon initial segment plays important roles in spike initiation and invasion of axonal spikes into the soma. Among primary sensory neurons, those in the mesencephalic trigeminal nucleus (MTN) are exceptional in their ability to initiate soma spikes (S-spikes) in response to synaptic inputs, consequently displaying two kinds of S-spikes, one caused by invasion of an axonal spike arising from the sensory receptor and the other initiated by somatic inputs. We investigated where spikes are initiated in such MTN neurons and whether there are any differences between the two kinds of S-spikes. Simultaneous patch-clamp recordings from the soma and axon hillock revealed a spike-backpropagation from the spike-initiation site in the stem axon to the soma in response to 1-ms somatic current pulse, which disclosed the delayed emergence of S-spikes after the current-pulse offset. These initiated S-spikes were smaller in amplitude than S-spikes generated by stimulation of the stem axon; however, 4-AP (≤0.5 mM) eliminated the amplitude difference. Furthermore, 4-AP dramatically shortened the delay in spike initiation without affecting the spike-backpropagation time in the stem axon, whereas it substantially prolonged the refractory period of S-spikes arising from axonal-spike invasion without significantly affecting that of presumed axonal spikes. These observations suggest that 4-AP–sensitive K⁺ currents exert two opposing effects on S-spikes depending on their origins: suppression of spike initiation and facilitation of axonal-spike invasion at higher frequencies. Consistent with these findings, strong immunoreactivities for Kv1.1 and Kv1.6, among 4-AP–sensitive and low-voltage–activated Kv1 family examined, were detected in the soma but not in the stem axon of MTN neurons.

The Na⁺–K⁺pump current (I_p) and the h-current (I_h) flowing through hyperpolarization-activated channels (h-channels) participate in generating the resting potential. These two currents are thought to be produced independently. We show here bidirectional interactions between Na⁺–K⁺pumps and h-channels in mesencephalic trigeminal neurons. Activation ofI_hleads to the generation of two types of ouabain-sensitiveI_pwith temporal profiles similar to those of instantaneous and slow components ofI_h, presumably reflecting Na⁺transients in a restricted cellular space. Moreover, theI_pactivated by instantaneousI_hcan facilitate the subsequent activation of slowI_h. Such counteractive and cooperative interactions were also disclosed by replacing extracellular Na⁺with Li⁺, which is permeant through h-channels but does not stimulate the Na⁺–K⁺pump as strongly as Na⁺ions. These observations indicate that the interactions are bidirectional and mediated by Na⁺ions. Also after substitution of extracellular Na⁺with Li⁺, the tailI_hwas reduced markedly despite an enhancement ofI_hitself, attributable to a negative shift of the reversal potential forI_hpresumably caused by intracellular accumulation of Li⁺ions. This suggests the presence of a microdomain where the interactions can take place. Thus, the bidirectional interactions between Na⁺–K⁺pumps and h-channels are likely to be mediated by Na⁺microdomain. Consistent with these findings, hyperpolarization-activated and cyclic nucleotide-modulated subunits (HCN1/2) and the Na⁺–K⁺pumpα3 isoform were colocalized in plasma membrane of mesencephalic trigeminal neurons having numerous spines.

Learning and memory are critically dependent on basal forebrain cholinergic (BFC) neuron excitability, which is modulated profoundly by leak K⁺ channels. Many neuromodulators closing leak K⁺ channels have been reported, whereas their endogenous opener remained unknown. We here demonstrate that nitric oxide (NO) can be the endogenous opener of leak K⁺ channels in the presumed BFC neurons. Bath application of 1 mM S-nitroso- N-acetylpenicillamine (SNAP), an NO donor, induced a long-lasting hyperpolarization, which was often interrupted by a transient depolarization. Soluble guanylyl cyclase inhibitors prevented SNAP from inducing hyperpolarization but allowed SNAP to cause depolarization, whereas bath application of 0.2 mM 8-bromoguanosine-3′,5′-cyclomonophosphate (8-Br-cGMP) induced a similar long-lasting hyperpolarization alone. These observations indicate that the SNAP-induced hyperpolarization and depolarization are mediated by the cGMP-dependent and -independent processes, respectively. When examined with the ramp command pulse applied at –70 mV under the voltage-clamp condition, 8-Br-cGMP application induced the outward current that reversed at K⁺ equilibrium potential ( E_K) and displayed Goldman-Hodgkin-Katz rectification, indicating the involvement of voltage-independent K⁺ current. By contrast, SNAP application in the presumed BFC neurons either dialyzed with the GTP-free internal solution or in the presence of 10 μM Rp-8-bromo-β-phenyl-1,N²-ethenoguanosine 3′,5′-cyclic monophosphorothioate sodium salt, a protein kinase G (PKG) inhibitor, induced the inward current that reversed at potentials much more negative than E_K and close to the reversal potential of Na⁺-K⁺ pump current. These observations strongly suggest that NO activates leak K⁺ channels through cGMP-PKG-dependent pathway to markedly decrease the excitability in BFC neurons, while NO simultaneously causes depolarization by the inhibition of Na⁺-K⁺ pump through ATP depletion.

Leak K⁺ conductance generated by TASK1/3 channels is crucial for neuronal excitability. However, endogenous modulators activating TASK channels in neurons remained unknown. We previously reported that in the presumed cholinergic neurons of the basal forebrain (BF), activation of NO-cGMP-PKG (protein kinase G) pathway enhanced the TASK1-like leak K⁺ current (I-K_leak). As 8-Br-cGMP enhanced the I-K_leak mainly at pH 7.3 as if changing the I-K_leak from TASK1-like to TASK3-like current, such an enhancement of the I-K_leak would result either from an enhancement of hidden TASK3 component or from an acidic shift in the pH sensitivity profile of TASK1 component. In view of the report that protonation of TASK channel decreases its open probability, the present study was designed to examine whether the activation of PKG increases the conductance of TASK1 channels by reducing their binding affinity for H⁺, i.e., by increasing K_d for protonation, or not. We here demonstrate that PKG activation and inhibition respectively upregulate and downregulate TASK1 channels heterologously expressed in PKG-loaded HEK293 cells at physiological pH, by causing shifts in the K_d in the acidic and basic directions, respectively. Such PKG modulations of TASK1 channels were largely abolished by mutating pH sensor H98. In the BF neurons that were identified to express ChAT and TASK1 channels, similar dynamic modulations of TASK1-like pH sensitivity of I-K_leak were caused by PKG. It is strongly suggested that PKG activation and inhibition dynamically modulate TASK1 currents at physiological pH by bidirectionally changing K_d values for protonation of the extracellular pH sensors of TASK1 channels in cholinergic BF neurons.