In Vitro Electrophysiological Recordings From Brain Slices

Recordings of neurons maintained in vitro have led to significant advances in understanding the ionic mechanisms underlying neurotransmitter and drug actions. This preparation consists of slices 300-400 Mm thick cut from the brain of an animal soon after decapitation. If this procedure is done carefully and the brain slices are rapidly placed into oxygenated physiological saline, the neurons within the slices will remain alive and healthy, often for 10 hours or more. Because the neurons are recorded in a chamber with oxygenated media superfused over the slice, several advantages may be realized:

1. Both intracellular and extracellular recordings are more stable because blood and breathing pulsations are absent.

2. Visual control over electrode placement is achieved.

3. The ionic composition of the microenvironment may be controlled precisely.

4. Little interference from the activity of long-loop afferents occurs, and the near-absence of spontaneous spike discharge limits the contribution of local circuit neurons to the responses.

Furthermore, in contrast to microiontophoresis, the concentration of drug in the solution can be controlled precisely. This preparation is also the most complex that can be used for patch clamp recordings because debris may be removed and the patch pipette placed on selected neurons under visual control with a high-resolution optics system (Edwards et al. 1989).

Nonetheless, because of the isolated nature of this system, the results obtained may not precisely reflect the physiology of the intact system. For example, dopamine neurons recorded in vivo have been characterized by their burst-firing discharge pattern (Grace and Bunney 1984a), which appears to be important in regulating neurotransmitter release (Gonon 1988). However, dopamine neurons recorded in vitro do not fire in bursting patterns (Grace and Onn 1989) (Figure 5-9). On the other hand, this distinction provides what may be an ideal system for examining the factors that cause in vivo burst firing. Therefore, the most complete model of the functioning of a system or of its response to drug application can be derived by comparing the results obtained in vitro with those in the intact organism.

FIGURE 5-9. Variation (sometimes substantial) in patterns of activity of a neuron type, depending on the preparation in which it is recorded.

(A) Extracellular recordings of a dopamine neuron in an intact anesthetized rat (i.e., in vivo) illustrate the typical irregular firing pattern of the cell, with single spikes occurring intermixed with bursts of action potentials. (B) In contrast, intracellular recordings of a dopamine neuron in an isolated brain slice preparation (i.e., in vitro) illustrate the pacemaker pattern that occurs exclusively in identified dopamine neurons in this preparation. For dopamine neurons, a pacemaker firing pattern is rarely observed in vivo, and burst firing has never been observed in the in vitro preparation. However, although the activity recorded in vitro is obviously an abstraction compared with the firing pattern of this neuron in vivo, a comparative study in each preparation does provide the opportunity to examine factors that may underlie the modulation of firing pattern in this neuronal type.

Source. Adapted from Grace AA: "The Regulation of Dopamine Neuron Activity as Determined by In Vivo and In Vitro Intracellular Recordings," in The Neurophysiology of Dopamine Systems. Edited by Chiodo LA, Freeman AS. Detroit, MI, Lake Shore Publications, 1987, pp. 1-67 (Copyright 1987, Lake Shore Publications. Used with permission); and Grace AA, Bunney BS: "Intracellular and Extracellular Electrophysiology of Nigral Dopaminergic Neurons, I: Identification and Characterization." Neuroscience 10:301-315, 1983. Copyright 1983, International Brain Research Organization. Used with permission.

Recordings From Dissociated Neurons and Neuronal Cell Cultures

Recordings from isolated neurons are actually a subset of in vitro recording methods, with many of the same advantages in terms of accessibility and stability. Furthermore, because the neurons can be completely visualized, advanced techniques such as patch clamping are more easily done. A unique advantage of this system can be obtained by coculturing different neuronal populations. For example, defining the effects of a noradrenergic synapse on a hippocampal pyramidal neuron more precisely may be possible by coculturing these cell types and allowing them to make synapses. In this way, the researcher has visual control over impaling neurons that constitute a presynaptic and postsynaptic pairing. On the other hand, the synapses formed are not necessarily limited to those that occur naturally in the intact organism, in terms of both the location of the synapse on the neuron and the classes of neurons that are interconnected. Furthermore, the altered neuronal morphology present in these preparations may modify the response of the neurons to drugs. Nonetheless, when the analysis is limited to well-defined responses, such as second-messenger actions or ion channel measures, this system affords an unparalleled level of accessibility.

Brain Blaster

Brain Blaster

Have you ever been envious of people who seem to have no end of clever ideas, who are able to think quickly in any situation, or who seem to have flawless memories? Could it be that they're just born smarter or quicker than the rest of us? Or are there some secrets that they might know that we don't?

Get My Free Ebook


Post a comment