Temporal coupling between neuronal activity and blood flow in rat cerebellar cortex as indicated by field potential analysis

Abstract

1. Laser-Doppler flowmetry and extracellular recordings of field potentials were used to examine the temporal coupling between neuronal activity and increases in cerebellar blood flow (CeBF). 2. Climbing fibre-evoked increases in CeBF were dependent on stimulus duration, indicating that increases in CeBF reflected a time integral in neuronal activity. The simplest way to represent neuronal activity over time was to obtain a running summation of evoked field potential amplitudes (runSigmaFP). RunSigmaFP was calculated for each stimulus protocol and compared with the time course of the CeBF responses to demonstrate coupling between nerve cell activity and CeBF. 3. In the climbing fibre system, the amplitude and time course of CeBF were in agreement with the calculated postsynaptic runSigmaFP (2-20 Hz for 60 s). This suggested coupling between CeBF and neuronal activity in this excitatory, monosynaptic, afferent-input system under these conditions. There was no correlation between runSigmaFP and CeBF during prolonged stimulation. 4. Parallel fibre-evoked increases in CeBF correlated with runSigmaFP of pre- and postsynaptic potentials (2-15 Hz for 60 s). At higher stimulation frequencies and during longer-lasting stimulation the time course and amplitudes of CeBF responses correlated with runSigmaFP of presynaptic, but not postsynaptic potentials. This suggested a more complex relationship in this mixed inhibitory-excitatory, disynaptic, afferent-input system. 5. This study has demonstrated temporal coupling between neuronal activity and CeBF in the monosynaptic, excitatory climbing-fibre system. In the mixed mono- and disynaptic parallel fibre system, temporal coupling was most clearly observed at low stimulation frequencies. We propose that appropriate modelling of electrophysiological data is needed to document functional coupling of neuronal activity and blood flow.

Figure 1

Three-dimensional drawing showing laser-Doppler (LDF) probe, stimulating and recording electrodes, and the functional anatomy of the rat cerebellar cortex including the molecular layer (Mol, thickness 200–400 μm), the Purkinje cell layer (PcL, thickness of about 100 μm), and the granular cell layer (GrL, thickness 400–500 μm), and the white matter below. The molecular layer contains granule cell axons, i.e. parallel fibres (PF, running perpendicular to the sagittal plane), dendrites of Purkinje cells (flattened so that they lie parallel to the sagittal plane), and interneurones (B, basket cells; S, stellate cells; GC, Golgi cells). The granule cell layer (GrL) consists primarily of granule cells (GrC) which receive synaptic input from mossy fibres (MF). The superficial parallel fibres were stimulated by a bipolar stimulating electrode, while climbing fibres (CFs) were stimulated by a mono-polar electrode lowered into the caudal part of the inferior olive (IO), which projects to the lobule V and VI of the vermis region. Field potentials were recorded by a glass microelectrode. CeBF was recorded by a LDF-probe located 0.3–0.5 mm above the pial (Pia) surface, using green laser light and near-infrared laser light.

Figure 2. Field potentials recorded in the cerebellar cortex

A, climbing fibre evoked field potentials (five single sweeps) recorded at a depth of 300 μm. The arrow indicates the amplitude of the excitatory postsynaptic potential (EPSP). B, parallel fibre evoked field potentials (five single sweeps), recorded at a depth of 200 μm, consisting of a presynaptic potential marked PSP and an excitatory postsynaptic potential marked EPSP. The amplitudes of the PSP and the EPSP are marked by the double-headed arrows.

Figure 3. Calculation of running summation field potential amplitudes (runΣFPs)

The graph shows field potential amplitude values (FP, -[cross]-) and the running summations over a period of five responses (runΣFP, -○-). The first FP values were 0, 0, 0, 0, 0, 8, 7, 6, 5. The corresponding runΣFP values were 0, 8, 15, 21, 26. Thus, summation over five field potential values (FPs) led to a peak shift corresponding to the duration of the run summation period, which in this example corresponded to five stimulus-evoked responses.

Figure 4. Stimulus-train duration characteristics of CeBF increases in response to climbing fibre stimulation at 5 Hz

A, original CeBF trace from one rat in response to increasing duration of stimulus periods from 10 to 300 s. Horizontal bars at bottom indicate the duration of climbing fibre activation. B, maximal increases in CeBF as a function of stimulus periods of 10, 30, 60, 120, 300 and 600 s (n = 5).

Figure 5. Comparison of climbing fibre-evoked increases in CeBF and evoked field potential amplitudes during increasing running summation periods

A, increases in CeBF during 5 Hz stimulation for 60 s and the corresponding evoked field potential amplitudes (B). RunΣFPs summated over 10 s (C), over 20 s (D), and over 30 s (E). Increases in CeBF plotted against field potential amplitudes (FP, F), and against runΣFPs summated over periods of 10 s (G), 20 s (H) and 30 s (I). The maximal correlation coefficient was obtained using run summation periods of 20 s (r = 0.902).

Figure 6. Climbing fibre evoked increases in CeBF compared with runΣFPs

Increases in CeBF and field potentials were evoked at 2, 5, 7, 10, 15 and 20 Hz stimulation for 60 s and by prolonged stimulation at 5 Hz for 600 s. CeBF increased frequency dependently in the range from 2 to 10 Hz and reached a plateau at 150 %. The CeBF elevation started to decrease before end of stimulus train at stimulus frequencies above 2 Hz. The runΣFPs were generated using periods of 20 s for the 2 to 15 Hz responses, and 30 s for the 20 Hz responses, and superimposed onto the CeBF trace. The ordinate for runΣFP traces were scaled to each other.

Figure 7. Comparison of parallel fibre-evoked increases in CeBF and evoked field potentials during increasing summation periods

A, increases in CeBF evoked at 10 and 20 Hz stimulation for 60 s. Stimulation periods are indicated by horizontal bars below the CeBF traces. B, the changes in CeBF were plotted against presynaptic field potential amplitudes using running summation periods for 40, 50 and 60 s. The maximal correlation coefficients were obtained using running summation periods of 60 s for the 10 Hz responses (r = 0.950) and 50 s for the 20 Hz responses (r = 0.941). C, the changes in CeBF were also plotted against postsynaptic runΣFPs summated over periods of 40 s, 50 s and 60 s. The maximal linear correlation coefficients were obtained using running summation periods of 60 s for the 10 Hz responses (r = 0.945). At 20 Hz stimulation, it was not possible to obtain a linear correlation between changes in CeBF and runΣFPs.

Figure 8. Increases in CeBF during parallel fibre stimulation

Increases in CeBF during parallel fibre stimulation compared with runΣFPs for the presynaptic potentials (thick continuous line) and the excitatory postsynaptic potentials (thin continuous line). Increases in CeBF were evoked by parallel fibre stimulation at 2–30 Hz for 60 s, and 10 Hz for 600 s. The presynaptic runΣFPs were summated over periods of 20 s at 2 Hz, 50 s at 5 Hz, 50–60 s at 10–15 Hz and 40 s at 20–30 Hz stimulation (thick continuous line). The postsynaptic runΣFPs were summated over periods of 20 s for the 2 Hz response and 50 s for 5–30 Hz response (thin continuous line). The ordinate for runΣFP were scaled to each other. The time course of CeBF was modelled by the presynaptic runΣFP at 2–20 Hz stimulation and during long-lasting stimulation (10 Hz for 600 s), while the postsynaptic runΣFP of the EPSP only mimicked the time course of the CeBF trace at low stimulation frequencies (2–15 Hz). The postsynaptic runΣFPs did not model changes in CeBF at 20 and 30 Hz stimulation. A similar pattern was observed during long-lasting stimulation (10 Hz for 600 s).