Loose coupling between SK and P/Q-type Ca2+ channels in cartwheel cells of the dorsal cochlear nucleus

Small-conductance Ca2+-activated K+ (SK) and large-conductance voltage- and Ca2+-activated K+ (BK) channels are Ca2+-activated K+ channels that control action potential firing in diverse neurons in the brain. In cartwheel cells of the dorsal cochlear nucleus, blockade of either channel type leads to excessive production of spike bursts. In the same cells, P/Q-type Ca2+ channels in plasma membrane and ryanodine receptors in endoplasmic reticulum supply Ca2+ to BK channels through Ca2+ nanodomain signaling. In this study, voltage-clamp experiments were performed in cartwheel cells in mouse brain slices to examine the Ca2+ signaling pathways underlying activation of SK channels. As with BK channels, SK channels required the activity of P/Q-type Ca2+ channels. However, this signaling occurred across Ca2+ micro- rather than nanodomain distances and was independent of Ca2+ release from endoplasmic reticulum. These differential modes of activation may lead to distinct time courses of the two K+ currents and therefore control excitability of auditory neurons across different timescales. NEW & NOTEWORTHY This study has shown for the first time that in cartwheel cells of the dorsal cochlear nucleus, small-conductance Ca2+-activated K+ (SK) channels were triggered by the activation of P/Q-type Ca2+ channels in which SK–P/Q-type coupling is mediated within the Ca2+ microdomains (loose coupling). Although Ca2+-induced Ca2+ release is able to activate large-conductance voltage- and Ca2+-activated K+ (BK) channels in cartwheel cells, it did not contribute to SK activation.

Molecular Layer Inhibitory Interneurons Provide Feedforward and Lateral Inhibition in the Dorsal Cochlear Nucleus

In the outer layers of the dorsal cochlear nucleus, a cerebellum-like structure in the auditory brain stem, multimodal sensory inputs drive parallel fibers to excite both principal (fusiform) cells and inhibitory cartwheel cells. Cartwheel cells, in turn, inhibit fusiform cells and other cartwheel cells. At the microcircuit level, it is unknown how these circuit components interact to modulate the activity of fusiform cells and thereby shape the processing of auditory information. Using a variety of approaches in mouse brain stem slices, we investigated the synaptic connectivity and synaptic strength among parallel fibers, cartwheel cells, and fusiform cells. In paired recordings of spontaneous and evoked activity, we found little overlap in parallel fiber input to neighboring neurons, and activation of multiple parallel fibers was required to evoke or alter action potential firing in cartwheel and fusiform cells. Thus neighboring neurons likely respond best to distinct subsets of sensory inputs. In contrast, there was significant overlap in inhibitory input to neighboring neurons. In recordings from synaptically coupled pairs, cartwheel cells had a high probability of synapsing onto nearby fusiform cells or other nearby cartwheel cells. Moreover, single cartwheel cells strongly inhibited spontaneous firing in single fusiform cells. These synaptic relationships suggest that the set of parallel fibers activated by a particular sensory stimulus determines whether cartwheel cells provide feedforward or lateral inhibition to their postsynaptic targets.

Two Distinct Types of Inhibition Mediated by Cartwheel Cells in the Dorsal Cochlear Nucleus

Individual neurons have been shown to exhibit target cell-specific synaptic function in several brain areas. The time course of the postsynaptic conductances (PSCs) strongly influences the dynamics of local neural networks. Cartwheel cells (CWCs) are the most numerous inhibitory interneurons in the dorsal cochlear nucleus (DCN). They are excited by parallel fiber synapses, which carry polysensory information, and in turn inhibit other CWCs and the main projection neurons of the DCN, pyramidal cells (PCs). CWCs have been implicated in “context-dependent” inhibition, producing either depolarizing (other CWCs) or hyperpolarizing (PCs) post synaptic potentials. In the present study, we used paired whole cell recordings to examine target-dependent inhibition from CWCs in neonatal rat DCN slices. We found that CWC inhibitory postsynaptic potentials (IPSPs) onto PCs are large (1.3 mV) and brief (half-width = 11.8 ms), whereas CWC IPSPs onto other CWCs are small (0.2 mV) and slow (half-width = 36.8 ms). Evoked IPSPs between CWCs exhibit paired-pulse facilitation, while CWC IPSPs onto PCs exhibit paired-pulse depression. Perforated-patch recordings showed that spontaneous IPSPs in CWCs are hyperpolarizing at rest with a mean estimated reversal potential of −67 mV. Spontaneous IPSCs were smaller and lasted longer in CWCs than in PCs, suggesting that the kinetics of the receptors are different in the two cell types. These results reveal that CWCs play a dual role in the DCN. The CWC-CWC network interactions are slow and sensitive to the average rate of CWC firing, whereas the CWC-PC network is fast and sensitive to transient changes in CWC firing.

Distinct Functional and Anatomical Architecture of the Endocannabinoid System in the Auditory Brainstem

Endocannabinoids (ECs) act as retrograde messengers that enable postsynaptic cells to regulate the strength of their synaptic inputs. Here, by using physiological and histological techniques, we showed that, unlike in other parts of the brain, excitatory inputs are more sensitive than inhibitory inputs to EC signaling in the dorsal cochlear nucleus (DCN), an auditory brainstem nucleus. The principal cells of the DCN, fusiform cells, integrate acoustic signals through nonplastic synapses located in the deep layer with multimodal sensory signals carried by plastic parallel fibers in the molecular layer. Parallel fibers contact fusiform cells and inhibitory interneurons, the cartwheel cells, which in turn inhibit fusiform cells. Postsynaptic depolarization or pairing of postsynaptic potentials (PSPs) with action potentials (APs) induced EC-mediated modulation of excitatory inputs but did not affect inhibitory inputs. Quantitative electron microscopical studies showed that glutamatergic terminals express more cannabinoid 1 receptors (CB1Rs) than glycinergic terminals. Fusiform and cartwheel cells express diacylglycerol lipase α and β (DGLα/β), the two enzymes involved in the generation of the EC, 2-arachidonoyl-glycerol (2-AG). DGLα and DGLβ are found in the spines of cartwheel but not fusiform cells indicating that the synthesis of ECs is more distant from parallel fiber synapses in fusiform than cartwheel cells. The differential localization and density of DGLα/β and CB1Rs leads to cell- and input-specific EC signaling that favors activity-dependent EC-mediated suppression at synapses between parallel fibers and cartwheel cell spines, thus leading to reduced feedforward inhibition in fusiform cells. We propose that EC signaling is a major modulator of the balance of excitation and inhibition in auditory circuits.

Temporal and Frequency Characteristics of Cartwheel Cells in the Dorsal Cochlear Nucleus of the Awake Mouse

The dorsal cochlear nucleus (DCN) is an initial site of central auditory processing and also the first site of multisensory convergence in the auditory pathway. The auditory nerve imparts a tonotopic frequency organization on the responses of principal cells in the DCN. Cartwheel cells modify the responses of principal cells, but they do not receive direct auditory nerve input. This study shows that cartwheel cells respond well to tonal stimuli in the awake mouse and they have a well-defined characteristic frequency that corresponds to the tonotopic organization of the DCN. The auditory responses of cartwheel cells exhibit complex spectrotemporal responses to tones, with excitation and inhibition modulating the firing patterns in both frequency and time after onset of the stimulus. Temporal responses to best-frequency tones are highly variable between cartwheel cells, but a simple model is used to unify this variability as differences in the timing of synaptic currents. Cartwheel cell responses to two-tone stimuli show that interactions from different frequencies affect the output of cartwheel cells. The results suggest that at this primary auditory structure, processing of sound at one frequency can be modified by sounds of different frequency. These complex frequency and temporal interactions in cartwheel cells suggest that these neurons play an active role in basic sound processing.

Ion Channels Generating Complex Spikes in Cartwheel Cells of the Dorsal Cochlear Nucleus

Cartwheel cells are glycinergic interneurons that modify somatosensory input to the dorsal cochlear nucleus. They are characterized by firing of mixtures of both simple and complex action potentials. To understand what ion channels determine the generation of these two types of spike waveforms, we recorded from cartwheel cells using the gramicidin perforated-patch technique in brain slices of mouse dorsal cochlear nucleus and applied channel-selective blockers. Complex spikes were distinguished by whether they arose directly from a negative membrane potential or later during a long depolarization. Ca2+ channels and Ca2+-dependent K+ channels were major determinants of complex spikes. Onset complex spikes required T-type and possibly R-type Ca2+ channels and were shaped by BK and SK K+ channels. Complex spikes arising later in a depolarization were dependent on P/Q- and L-type Ca2+ channels as well as BK and SK channels. BK channels also contributed to fast repolarization of simple spikes. Simple spikes featured an afterdepolarization that is probably the trigger for complex spiking and is shaped by T/R-type Ca2+ and SK channels. Fast spikes were dependent on Na+ channels; a large persistent Na+ current may provide a depolarizing drive for spontaneous activity in cartwheel cells. Thus the diverse electrical behavior of cartwheel cells is determined by the interaction of a wide variety of ion channels with a prominent role played by Ca2+.