sharp glass pipette electrodes  (Sutter Instrument Company)

Journal: The Journal of Neuroscience

Article Title: Respiratory and Telencephalic Modulation of Vocal Motor Neurons in the Zebra Finch

doi: 10.1523/JNEUROSCI.23-03-01072.2003

Figure Lengend Snippet: Morphological and intrinsic electrical features of XIIts MNs. A, Intracellular staining of an XIIts MN shows dendrites extending into regions surrounding nXIIts and an axon that travels ventrally along the midline toward the XIIth nerve rootlet. Motor neurons (gray polygons) in the rostral portion of XIIts were labeled by injection of retrograde tracer into the ventral syringeal muscle several days before intracellular filling in a brain slice preparation. D, Dorsal; L, lateral. B, In vitro, XIIts MNs generated highly regular trains of action potentials in response to positive currents. Membrane potential responses of an XIIts MN to positive and negative injected currents (±200 pA; the pulse duration is shown below the superimposed voltagetraces) passed through the recording electrode are shown from an in vitro intracellular recording made in a brain slice. The inset trace highlights the spike afterdepolarization (ADP) characteristic of XIIts MNs.C, An in vivo intracellular membrane potential recorded from a XIIts MN shows that in response to a +200 pA current injected through the recording electrode, the cell generates a train of action potentials similar to those seen in vitro, but with action potential suppression, likely to be occurring during the inspiratory phase of respiration (arrow) (see Fig. ​Fig.88 and Results). Note that subthreshold membrane potential movement during this inspiratory phase consists of a gradual rather than abrupt hyperpolarization. D, The average firing frequencies (in hertz; in vitro,left; in vivo, right) are plotted as a function of the amplitude of the positive current injected through the recording electrode for populations of XIIts MNs (in vitro, 40 cells; in vivo, 5 cells). The average firing frequency as a function of injected current in vitro and in vivo was highly linear between 0 and +0.6 nA. E, Instantaneous action potential frequency plots (in vitro, left; in vivo, right; averages from same cells as inD).

Article Snippet: Sharp electrodes (borosilicate glass, BF100–50-10, Sutter Instrument) were pulled to yield a resistance of 100–200 MΩ when filled with 2 m K-acetate and 5% neurobiotin.

Techniques: Staining, Labeling, Injection, Slice Preparation, In Vitro, Generated, Membrane, In Vivo

Journal: The Journal of Neuroscience

Article Title: Respiratory and Telencephalic Modulation of Vocal Motor Neurons in the Zebra Finch

doi: 10.1523/JNEUROSCI.23-03-01072.2003

Figure Lengend Snippet: In the presence of glycine receptor blockers, electrical stimulation in ipsilateral RAm evoked EPSPs in XIIts MNs mediated in part by NMDA receptors and which were sufficient to translate single EPSPs into a sustained action potential discharge.A, A single EPSP could trigger sustained trains of action potentials at the actual resting potential of the cell (actual Vm) but was rendered highly phasic and entirely subthreshold when the impaled XIIts MN was slightly hyperpolarized via tonic negative current injection through the recording electrode (more negative Vm). B, The slow portion of the EPSP evoked by RAm stimulation in brain slices bathed in strychnine had the voltage-dependence characteristic of NMDA receptor-mediated transmission and was strongly attenuated by hyperpolarization of the postsynaptic XIIts membrane. The EPSP evoked by RAm stimulation is displayed at the actual resting potential of the cell (topmost trace; actual Vm) or at a slightly more negative membrane potential (bottom trace;more negative Vm) achieved by passing tonic current through the recording electrode. The EPSP triggered a train of action potentials at the more positive membrane potential. Subsequent treatment with the NMDA receptor blocker d-APV reduced the EPSP in a manner resembling postsynaptic hyperpolarization, localizing the NMDA receptors to the impaled cell. C, The slow portion of the EPSP was blocked reversibly by applying NMDA receptor antagonists to XIIts. EPSPs that were collected before and during application of the NMDA receptor antagonist d-APV to XIIts are shown superimposed (see Materials and Methods). These results suggest that NMDA receptors on the XIIts MN are activated by lateral medullary inputs. Raw traces are shown in A andB, and averages of three to six traces are shown inC. Stim, Stimulation time.

Article Snippet: Sharp electrodes (borosilicate glass, BF100–50-10, Sutter Instrument) were pulled to yield a resistance of 100–200 MΩ when filled with 2 m K-acetate and 5% neurobiotin.

Techniques: Injection, Transmission Assay, Membrane

Journal: The Journal of Neuroscience

Article Title: Respiratory and Telencephalic Modulation of Vocal Motor Neurons in the Zebra Finch

doi: 10.1523/JNEUROSCI.23-03-01072.2003

Figure Lengend Snippet: Membrane potential manipulations reveal phasic excitation at expiratory EMG onset. An in vivointracellular recording (mV) from an XIIts MN and a simultaneous abdominal muscle EMG (EMG) were used to measure the nature of respiratory activity in XIIts. During these recordings, the XIIts MN membrane potential was either slightly depolarized (top) or moderately or more strongly hyperpolarized (middle and bottom) by injecting positive or negative currents through the recording electrode. When slightly depolarized from its resting membrane potential value with positive current (+0.5 nA, top), the neuron showed only gradual membrane hyperpolarization (but abrupt firing rate suppression) during the inspiratory phase. When moderately or more substantially hyperpolarized from its resting membrane potential with tonic negative current passed through the recording electrode (−1.25 nA, middle; −1.5 nA,bottom), the neuron displayed subthreshold excitation occurring abruptly at the onset of expiration. No membrane potential movements were noted at the offset of expiratory EMG activity.

Article Snippet: Sharp electrodes (borosilicate glass, BF100–50-10, Sutter Instrument) were pulled to yield a resistance of 100–200 MΩ when filled with 2 m K-acetate and 5% neurobiotin.

Techniques: Membrane, Activity Assay

Journal: The Journal of Neuroscience

Article Title: Respiratory and Telencephalic Modulation of Vocal Motor Neurons in the Zebra Finch

doi: 10.1523/JNEUROSCI.23-03-01072.2003

Figure Lengend Snippet: Changes in XIIts MN input impedance during BOS playback and during subsequent respiratory entrainment. Forward (bottom left) and reverse BOS (REV BOS) (bottom right) playback-evoked vocal and respiratory activity, as revealed by simultaneous abdominal expiratory EMGs (top) and intracellular recordings from an XIIts neuron [action potential PSTH (XIIts spikes) and median-filtered average membrane potential (XIIts V¯m)]. In the lower of the two median filtered traces, the XIIts MN was injected with brief (20 msec) hyperpolarizing (−0.5 nA) currents through the recording electrode at 8 Hz to monitor postsynaptic impedance changes indicative of changes in synaptic conductances. Forward BOS strongly excited the XIIts neuron, resulting in marked impedance decreases (open arrows), consistent with elevated excitatory synaptic drive during song playback. Note that expiratory activity increased just before the augmentation in XIIts MN (enhanced EMG activity is marked by a solid arrow). Inspiration, reflected as troughs in the EMG, consistently followed forward BOS playback in this case. Such respiratory entrainment also occurred in response to reverse BOS playback, although this stimulus did not excite the XIIts neuron (note different y-axis scales for the forward vs reverse BOS-evoked XIIts PSTHs). Postsynaptic impedance decreases, measured as decrements in the amplitude of the DC-evoked hyperpolarizations, were not detected during such entrained inspiration, although slight impedance decreases were noted during subsequent expiratory phases (marked by open arrows, aligned to XIIts spiking and EMGs with closed arrows). These results are consistent with the idea that the XIIts respiratory rhythm derives primarily or solely from expiratory-driven excitation. Seventy iterations each of forward and reverse BOS playback were used to construct the PSTHs of XIIts MN and expiratory EMG activity, whereas 20 iterations were used to generate each of the two median filteredVm values (i.e., current pulses were not injected through the recording electrode on every trial).

Article Snippet: Sharp electrodes (borosilicate glass, BF100–50-10, Sutter Instrument) were pulled to yield a resistance of 100–200 MΩ when filled with 2 m K-acetate and 5% neurobiotin.

Techniques: Activity Assay, Membrane, Injection, Construct