Serapio M. Baca

Cortical spreading depression and migraine

Cortical spreading depression (CSD), a slowly propagated wave of depolarization followed by suppression of brain activity, is a remarkably complex event that involves dramatic changes in neural and vascular function. Since its original description in the 1940s, CSD has been hypothesized to be the underlying mechanism of the migraine aura. Substantial evidence from animal models provides indirect support for this hypothesis, and studies showing that CSD is common in humans with brain injury clearly demonstrate that the phenomenon can occur in the human brain. Considerable uncertainty about the role of CSD in migraine remains, however, and key questions about how this event is initiated, how it spreads, and how it might cause migraine symptoms remain unanswered. This Review summarizes current concepts of CSD and its potential roles in migraine, and addresses ongoing studies aimed at a clearer understanding of this fundamental brain phenomenon.

Glycinergic Pacemaker Neurons in PreBötzinger Complex of Neonatal Mouse

The preBötzinger complex (preBötC) is essential for normal respiratory rhythm generation in rodents, for which the underlying mechanisms remain unknown. Excitatory preBötC pacemaker neurons are proposed to be necessary for rhythm generation. Here we report the presence of a population of preBötC glycinergic pacemaker neurons. We used rhythmic in vitro transverse slice preparations from transgenic mice where neurons expressing the glycine transporter 2 (GlyT2) gene coexpress enhanced green fluorescent protein (EGFP). We combined epifluorescence and whole-cell patch-clamp recording to study preBötC EGFP-labeled, i.e., glycinergic, inspiratory-modulated neurons with pacemaker properties. We defined glycinergic pacemaker neurons as those preBötC EGFP neurons that exhibited the following: (1) ectopic bursting in rhythmic slices when depolarized during their normally silent period and (2) bursting when depolarized in nonrhythmic slices (following AMPA receptor blockade). Forty-two percent of EGFP-labeled neurons were inspiratory (n = 48 of 115), of which 23% (n = 11 of 48 inspiratory; 10% of the total recorded) were pacemakers. We conclude that there is a population of preBötC inspiratory-modulated glycinergic, presumably inhibitory, pacemaker neurons that constitute a substantial fraction of all preBötC pacemaker neurons. These findings challenge contemporary models for respiratory rhythmogenesis that assume the excitatory nature of preBötC pacemaker neurons. Testable and nontrivial predictions of the functional role of excitatory and inhibitory pacemaker neurons need to be proposed and the necessary experiments performed.

Widespread Inhibition Proportional to Excitation Controls the Gain of a Leech Behavioral Circuit

Changing gain in a neuronal system has important functional consequences, but the underlying mechanisms have been elusive. Models have suggested a variety of neuronal and systems properties to accomplish gain control. Here, we show that the gain of the neuronal network underlying local bending behavior in leeches depends on widespread inhibition. Using behavioral analysis, intracellular recordings, and voltage-sensitive dye imaging, we compared the effects of blocking just the known lateral inhibition with blocking all GABAergic inhibition. This revealed an additional source of inhibition, which was widespread and increased in proportion to increasing stimulus intensity. In a model of the input/output functions of the three-layered local bending network, we showed that inhibiting all interneurons in proportion to the stimulus strength produces the experimentally observed change in gain. This relatively simple mechanism for controlling behavioral gain could be prevalent in vertebrate as well as invertebrate nervous systems.

Sequential Development of Electrical and Chemical Synaptic Connections Generates a Specific Behavioral Circuit in the Leech

Neuronal circuits form during embryonic life, even before synapses are completely mature. Developmental changes can be quantitative (e.g., connections become stronger and more reliable) or qualitative (e.g., synapses form, are lost, or switch from electrical to chemical or from excitatory to inhibitory). To explore how these synaptic events contribute to behavioral circuits, we have studied the formation of a circuit that produces local bending (LB) behavior in leech embryos. This circuit is composed of three layers of neurons: mechanosensory neurons, interneurons, and motor neurons. The only inhibition in this circuit is in the motor neuron layer; it allows the animal to contract on one side while relaxing the opposite side. LB develops in two stages: initially touching the body wall causes circumferential indentation (CI), an embryonic behavior in which contraction takes place around the whole perimeter of the segment touched; one or 2 d later, the same touch elicits adult-like LB. Application of bicuculline methiodide in embryos capable of LB switched the behavior back into CI, indicating that the development of GABAergic connections turns CI into LB. Using voltage-sensitive dyes and electrophysiological recordings, we found that electrical synapses were present early and produced CI. Inhibition appeared later, shaping the circuit that was already connected by electrical synapses and producing the adult behavior, LB.

Optogenetic induction of cortical spreading depression in anesthetized and freely behaving mice

Cortical spreading depression, which plays an important role in multiple neurological disorders, has been studied primarily with experimental models that use highly invasive methods. We developed a relatively non-invasive optogenetic model to induce cortical spreading depression by transcranial stimulation of channelrhodopsin-2 ion channels expressed in cortical layer 5 neurons. Light-evoked cortical spreading depression in anesthetized and freely behaving mice was studied with intracortical DC-potentials, multi-unit activity and/or non-invasive laser Doppler flowmetry, and optical intrinsic signal imaging. In anesthetized mice, cortical spreading depression induction thresholds and propagation rates were similar for invasive (DC-potential) and non-invasive (laser Doppler flowmetry) recording paradigms. Cortical spreading depression-related vascular and parenchymal optical intrinsic signal changes were similar to those evoked with KCl. In freely behaving mice, DC-potential and multi-unit activity recordings combined with laser Doppler flowmetry revealed cortical spreading depression characteristics comparable to those under anesthesia, except for a shorter cortical spreading depression duration. Cortical spreading depression resulted in a short increase followed by prolonged reduction of spontaneous active behavior. Motor function, as assessed by wire grip tests, was transiently and unilaterally suppressed following a cortical spreading depression. Optogenetic cortical spreading depression induction has significant advantages over current models in that multiple cortical spreading depression events can be elicited in a non-invasive and cell type-selective fashion.

Modulation of respiratory output by cervical epidural stimulation in the anesthetized mouse

Respiration is produced and controlled by well-characterized brain stem nuclei, but the contributions of spinal circuits to respiratory control and modulation remain under investigation. Many respiratory studies are conducted in in vitro preparations (e.g., brain stem slice) obtained from neonatal rodents. While informative, these studies do not fully recapitulate the complex afferent and efferent neural circuits that are likely to be involved in eupnea (i.e., quiet breathing). To begin to investigate spinal contributions to respiration, we electrically stimulated the cervical spinal cord during unassisted respiration in anesthetized, intact mice. Specifically, we used epidermal electrical stimulation at 20 Hz and varied current intensity to map changes in respiration. Stimulating at 1.5 mA at cervical level 3 (C3) consistently caused a significant increase in respiratory frequency compared with prestimulation baseline and when compared with sham stimulations. The increase in respiratory frequency persisted for several minutes after epidural stimulation ceased. There was no change in tidal volume, and the estimated minute ventilation was increased as a consequence of the increase in respiratory frequency. Sigh frequency also increased during epidural stimulation at C3. Neither the increase in respiratory frequency nor the increase in sighing were observed after stimulation at other dorsal cervical levels. These findings suggest that the spinal circuits involved in the modulation of eupnea and sighing may be preferentially activated by specific endogenous inputs. Moreover, the cervical spinal cord may play a role in respiratory modulation that affects both eupneic respiration and sigh production in intact, adult mice.

Neurovascular mechanisms of migraine and cluster headache

Vascular theories of migraine and cluster headache have dominated for many years the pathobiological concept of these disorders. This view is supported by observations that trigeminal activation induces a vascular response and that several vasodilating molecules trigger acute attacks of migraine and cluster headache in susceptible individuals. Over the past 30 years, this rationale has been questioned as it became clear that the actions of some of these molecules, in particular, calcitonin gene-related peptide and pituitary adenylate cyclase-activating peptide, extend far beyond the vasoactive effects, as they possess the ability to modulate nociceptive neuronal activity in several key regions of the trigeminovascular system. These findings have shifted our understanding of these disorders to a primarily neuronal origin with the vascular manifestations being the consequence rather than the origin of trigeminal activation. Nevertheless, the neurovascular component, or coupling, seems to be far more complex than initially thought, being involved in several accompanying features. The review will discuss in detail the anatomical basis and the functional role of the neurovascular mechanisms relevant to migraine and cluster headache.

EHMTI-0244. Optogenetic elicitation of cortical spreading depression in unanesthetized, head-restrained mice

We tested the hypothesis that cortical spreading depression (CSD) could be initiated by optogenetically activating astrocytes in unanesthetized mice. Previously, we demonstrated that depolarizing astrocytes expressing the light-responsive channelrhodopsin-2 receptor (ChR2) elicited CSD in brain slices and in anesthetized mice.