Dual Gamma Rhythm Generators Control Interlaminar Synchrony in Auditory Cortex

Pharmacological manipulation reveals different mechanisms underlying the slow and fast gamma rhythms. A, Proportional change in mean (n = 5 slices from 5 rats) gamma power for slow, layer 2/3 gamma (30–45 Hz, gray bars) and fast, layer 4 gamma (50–80 Hz, black bars), induced by 800 nm kainate. Both rhythms were equally sensitive to GABA_A receptor blockade (gabazine, 1 μm). Layer 4, high gamma was significantly less sensitive to AMPA receptor blockade (SYM2206, 10 μm) and nonspecific gap junction conductance decrease [carbenoxolone (cbx), 0.2 mm[rsqb]. Layer 2/3, slow gamma was insensitive to NR2C/D-containing NMDAR blockade (PPDA, 10 μm), whereas this manipulation almost abolished layer 4, fast gamma. Asterisks indicate significantly different effects of each drug on the two gamma rhythms (p < 0.05, n = 5 slices from 5 rats). B, Example pooled power spectra (n = 5 slices from 5 rats) for layer 4 population responses to 800 nm kainate showing the differential effects of PPDA (left graph) and carbenoxolone (right graph).

Lamina-specific interneuron input/output behavior before and after frequency bifurcation. A, Example layer 2/3 FS interneuron recordings during exposure to 400 nm kainate (low gamma only) and 800 nm kainate (low and high gamma). Top traces show behavior at resting membrane potential (RMP) in each condition (−55 mV, low gamma only; −53 mV, low and high gamma). Bottom traces show EPSP inputs recorded from −70 mV in each case. Note the maintenance of low gamma frequency spike generation despite the addition of faster, smaller EPSPs in the low and high gamma condition. B, Example recordings from a layer 4 FS interneuron during 400 and 800 nm kainate application, RMPs of −57 and −52 mV, respectively. Bottom traces show EPSP inputs at −70 mV. Note the paucity of spike generation and EPSP inputs in the low gamma condition is transformed into spiking at high gamma frequencies, with high gamma frequency EPSP inputs in 800 nm kainate. C, Example recordings from layer 5 FS interneurons in the two conditions. RMPs were −56 and −54 mV. Note spike rates switch from low to high gamma frequency corresponding to an increase in rate of EPSPs from low to high gamma frequency. Calibration: 100 ms, 40 mV (RMP), 10 mV (EPSP).

Lamina-specific principal cell input/output behavior before and after frequency bifurcation. A, Example layer 2/3 regular spiking pyramidal cell recordings during exposure to 400 nm kainate (low gamma only) and 800 nm kainate (low and high gamma). Top traces show behavior at RMP in each condition (−63 mV, low gamma only; −66 mV, low and high gamma). Bottom traces show IPSP inputs recorded from −30 mV in each case. Note the maintenance of very sparse spike generation despite the addition of faster, smaller IPSPs in the low and high gamma condition. B, Example recordings from a layer 4 accommodating principal cell during 400 and 800 nm kainate application, with RMPs of −64 and −57 mV, respectively. Bottom traces show IPSP inputs at −70 mV. Note the paucity of spike generation accompanying low gamma frequency, relatively broad IPSPs in the low gamma condition is transformed into near 1:1 spiking on each period of the high gamma local field potential. Note also the marked increase in mean IPSP amplitude in 800 nm kainate. C, Example recordings from a layer 5 intrinsically bursting pyramidal cell in the two conditions. RMPs were −67 and −63 mV. Note that spike rates switch from sparse, with bursts, to a missed-beat pattern during 800 nm kainate. Note also the increase in rate of IPSPs in this cell type. Calibration: 100 ms, 30 mV (RMP), 8 mV (IPSP).

Higher gamma frequencies associate with faster pyramidal neuron IPSPs in layers 4 and 5 but not layer 2/3. Graph shows mean decay constants (single-exponential fit) for IPSPs (average of 50 IPSPs per principal cell subtype averaged over n = 5 cells from 5 slices in each case) recorded from principal cells in each layer held at −30 mV by depolarizing current injection. Black bars show data in the presence of 400 nm kainate (low-frequency gamma only). Gray bars show data in the presence of 800 nm kainate (low- and higher-frequency gamma rhythms coexpressed). *p < 0.05. Right shows example mean IPSPs from single neurons (IPSP amplitude maximum-referenced average of 50 events in each case). Black traces are taken in the presence of 400 nm kainate, gray traces in 800 nm. Calibration: 20 ms.

Computational model demonstrates laminar-specific frequency sensitivity with change in drive. A, Behavior of the model network in conditions mimicking low glutamatergic excitation (400 nm kainate). The model robustly reproduces the core features of the experimental data: a low-frequency gamma rhythm in layers 2/3 projected to layers 4 and 5; sparse spiking in principal cells; only interneuron recruitment in layers 2/3 and 5, abolished by uncoupling layers (right) (compare Figs. 3, 4). B, Elevated tonic drive to layer 4 principal cells generates a faster rhythm in the model granular layer accompanied by local, intense principal cell spiking. Note that the lower-frequency, layer 2/3 rhythm persists in the presence of this faster rhythm but at marginally lower power, but the layer 5 rhythm switches to the layer 4 input frequency. Each of these two effects is abolished with removal of interlaminar connections. Note the separate calibration bar (in red, right of PSD graphs) for power spectra from layer 4. Ci, Diagram showing the structure of the model (for details, see Materials and Methods). Cii, The frequency bifurcation is enhanced by interlaminar interactions. Model predicts the reduction in frequency seen at 800 nm kainate is in part generated by ascending inhibition onto superficial interneurons (I_g–I_s removed, gray spectrum). Ciii, Model shows that effects of PPDA can be attributed to reduction in E_g–E_g recurrent excitation alone. Figure shows simulated field potentials from layer 4 in the presence (black) and absence (gray) of E_g–E_g with NMDAR-like kinetics. Calibration: 0.2 V, 100 ms. LFP, Local field potential.

Layers 4 and 5 but not layer 2/3 principal cells phase lock to the faster gamma rhythm. A, Graphs showing mean (n = 5 cells from 5 slices each from a separate rat) spike probability for layer 2/3 (top), layer 4 (middle), and layer 5 (bottom) principal cells. Data are plotted as probability of observing a spike on any given gamma period relative to local field potential phase in 1 ms bins, with 0 ms being the peak negativity in the field on each period. Data are overlaid from the two kainate (KA) concentrations: 400 nm generating only the lower gamma frequency (gray line) and 800 nm generating both fast and slow gamma rhythms concurrently (black line). Mean example traces below show 100 ms epochs of local field potential averaged to each spike in the corresponding intracellular recording. Note the different probability values on the axes for each of the three principal cell types. B, Period-by-period analysis of the relationship between instantaneous layer 4 local field potential frequency and corresponding layer 4 principal cell spiking. Data are pooled for both 400 and 800 nm kainate conditions. Note the near 1:1 relationship between layer 4 principal cell spike rate and field potential frequency for gamma frequencies over ∼45 Hz.