Journal of Neurophysiology

Ozaita, A.; Martone, M. E.; Ellisman, M. H.; Rudy, B.

doi: N/Apmid: 12091563

Abstract Voltage-gated K + channels containing pore-forming subunits of the Kv3 subfamily have specific roles in the fast repolarization of action potentials and enable neurons to fire repetitively at high frequencies. Each of the four known Kv3 genes encode multiple products by alternative splicing of 3′ ends resulting in the expression of K + channel subunits differing only in their C-terminal sequence. The alternative splicing does not affect the electrophysiological properties of the channels, and its physiological role is unknown. It has been proposed that one of the functions of the alternative splicing of Kv3 genes is to produce subunit isoforms with differential subcellular membrane localizations in neurons and differential modulation by signaling pathways. We investigated the role of the alternative splicing of Kv3 subunits in subcellular localization by examining the brain distribution of the two alternatively spliced versions of the Kv3.1 gene (Kv3.1a and Kv3.1b) with antibodies specific for the alternative spliced C-termini. Kv3.1b proteins were prominently expressed in the somatic and proximal dendritic membrane of specific neuronal populations in the mouse brain. The axons of most of these neurons also expressed Kv3.1b protein. In contrast, Kv3.1a proteins were prominently expressed in the axons of some of the same neuronal populations, but there was little to no Kv3.1a protein expression in somatodendritic membrane. Exceptions to this pattern were seen in two neuronal populations with unusual targeting of axonal proteins, mitral cells of the olfactory bulb, and mesencephalic trigeminal neurons, which expressed Kv3.1a protein in dendritic and somatic membrane, respectively. The results support the hypothesis that the alternative spliced C-termini of Kv3 subunits regulate their subcellular targeting in neurons. Footnotes Address for reprint requests: B. Rudy, Dept. of Physiology and Neuroscience, New York University School of Medicine, 550 First Ave., New York, NY 10016 (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Cangiano, Lorenzo; Wallén, Peter; Grillner, Sten

doi: N/Apmid: 12091554

Abstract Single motoneurons and pairs of a presynaptic reticulospinal axon and a postsynaptic motoneuron were recorded in the isolated lamprey spinal cord, to investigate the role of calcium-dependent K + channels (K Ca ) during the afterhyperpolarization following the action potential (AHP), and glutamatergic synaptic transmission on the dendritic level. The AHP consists of a fast phase due to transient K + channels (fAHP) and a slower phase lasting 100–200 ms (sAHP), being the main determinant of spike frequency regulation. We now present evidence that the sAHP has two components. The larger part, around 80%, is abolished by superfusion of Cd 2+ (blocker of voltage-dependent Ca 2+ channels), by intracellular injection of 1,2-bis-( 2 -aminophenoxy)-ethane- N,N,N′,N′ -tetraacetic acid (BAPTA; fast Ca 2+ chelator), and by apamin (selective toxin for K Ca channels of the SK subtype). While 80% of the sAHP is thus due to K Ca channels, the remaining 20% is not mediated by Ca 2+ , either entering through voltage-dependent Ca 2+ channels or released from intracellular Ca 2+ stores. This Ca 2+ -independent sAHP component has a similar time course as the K Ca portion and is not due to a Cl − conductance. It may be caused by Na + -activated K + channels. Glutamatergic excitatory postsynaptic potentials (EPSPs) evoked by single reticulospinal axons give rise to a local Ca 2+ increase in the postsynaptic dendrite, mediated in part by N -methyl- d -aspartate (NMDA) receptors. The Ca 2+ levels remain elevated for several hundred milliseconds and could be expected to activate K Ca channels. If so, this activation should cause a local conductance increase in the dendrite that would shunt EPSPs following the first EPSP in a spike train. We have tested this in reticulospinal/motoneuronal pairs, by stimulating the presynaptic axon with spike trains at different frequencies. We compared the first EPSP and the following EPSPs in the control and after blockade with apamin. No difference was observed in EPSP amplitude or shape before and after apamin, either in normal Ringer or in Mg 2+ -free Ringer removing the voltage-dependent block of NMDA receptors. In conclusion, the local Ca 2+ entry during reticulospinal EPSPs does not cause an activation of K Ca channels sufficient to affect the efficacy of synaptic transmission. Thus the integration of synaptic signals at the dendritic level in motoneurons appears simpler than would otherwise have been the case. Footnotes Address for reprint requests: S. Grillner, Dept. of Neuroscience, Karolinska Institute, 17177 Stockholm, Sweden (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Priebe, Nicholas J.; Lisberger, Stephen G.

doi: N/Apmid: 12091561

Abstract Neurons in area MT, a motion-sensitive area of extrastriate cortex, respond to a step of target velocity with a transient-sustained firing pattern. The transition from a high initial firing rate to a lower sustained rate occurs over a time course of 20–80 ms and is considered a form of short-term adaptation. In the present paper, we compared the tuning of the adaptation to the neuron's tuning to direction and speed. The tuning of adaptation was measured with a condition/test paradigm in which a testing motion of the preferred direction and speed of the neuron under study was preceded by a conditioning motion: the direction and speed of the conditioning motion were varied systematically. The response to the test motion depended strongly on the direction of the conditioning motion. It was suppressed in almost all neurons by conditioning motion in the same direction and could be either suppressed or enhanced by conditioning motion in the opposite direction. Even in neurons that showed suppression for target motion in the nonpreferred direction, the adaptation and response direction tuning were the same. The speed tuning of adaptation was linked much less tightly to the speed tuning of the response of the neuron under study. For just more than 50% of neurons, the preferred speed of adaptation was more than 1 log unit different from the preferred response speed. Many neurons responded best when slow motions were followed by faster motions (acceleration) or vice versa (deceleration), suggesting that MT neurons may encode information about the change of target velocity over time. Finally, adaptation by conditioning motions of different directions, but not different speeds, altered the latency of the response to the test motion. The adaptation of latency recovered with shorter intervals between the conditioning and test motions than did the adaptation of response size, suggesting that latency and amplitude adaptation are mediated by separate mechanisms. Taken together with the companion paper, our data suggest that short-term motion adaptation in MT is a consequence of the neural circuit in MT and is not mediated by either input-specific mechanisms or intrinsic mechanisms related to the spiking of individual neurons. The circuit responsible for adaptation is tuned for both speed and direction and has the same direction tuning as the circuit responsible for the initial response of MT neurons. Footnotes Address for reprint requests: N. J. Priebe, Department of Neurobiology and Physiology, Northwestern University, 2153 North Campus Drive, Evanston, IL 60208 (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Muir, G. D.; Chu, T. K.

doi: N/Apmid: 12091537

Abstract We have previously demonstrated that, even though chicks are very precocial and can locomote within hours of hatching, they require a period of time to develop a mature stable walk. As an example, 1- to 2-day-old animals move with disproportionately small stride lengths compared with 10- to 14-day-old animals. The purpose of this study was to determine whether the maturation of walking, including the development of a mature stride length, depends on locomotor experience. We also investigated the development and experience-dependence nature of head bobbing, an optokinetic behavior that occurs during walking in birds. Chicks were randomly assigned to one of three groups receiving either increased locomotor experience (i.e., treadmill exercise), decreased locomotor experience (i.e., decreased housing space), or no alteration in locomotor experience. To assess the dependence of locomotor maturation on N -methyl- d -aspartate (NMDA)–type glutamate receptors, animals in each group were either given an NMDA antagonist (MK-801, 1 mg/kg intramuscularly daily) or saline control. Locomotor characteristics (stride length, leg support durations, horizontal head excursions) were quantified from videotaped recordings of chicks walking overground unrestrained on posthatching days 1 , 2, 4, 6, 8, and 10 . Animals subject to exercise restriction for at least 6 days moved with shortened stride lengths compared with age-matched treadmill-exercised or control animals, a change that was maintained for the duration of the study. NMDA antagonism also resulted in shortened stride lengths. Head bobbing behavior matured during the same posthatching time period. The rate of this maturation was also decreased by exercise restriction. Thus locomotor experience is required for normal development of locomotor behavior, even in very precocial animals. These results are discussed in terms of the possible neuroanatomical and neurophysiological mechanisms underlying experience- and activity-dependent changes during motor development. Footnotes Address for reprint requests: G. D. Muir, Biomedical Sciences, Western College of Veterinary Medicine, University of Saskatchewan, 52 Campus Dr., Saskatoon, SK S7N 5B4, Canada (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Ireland, David R.; Abraham, Wickliffe C.

doi: N/Apmid: 12091536

Abstract Previous studies have implicated phospholipase C (PLC)-linked Group I metabotropic glutamate receptors (mGluRs) in regulating the excitability of hippocampal CA1 pyramidal neurons. We used intracellular recordings from rat hippocampal slices and specific antagonists to examine in more detail the mGluR receptor subtypes and signal transduction mechanisms underlying this effect. Application of the Group I mGluR agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) suppressed slow- and medium-duration afterhyperpolarizations (s- and mAHP) and caused a consequent increase in cell excitability as well as a depolarization of the membrane and an increase in input resistance. Interestingly, with the exception of the suppression of the mAHP, these effects were persistent, and in the case of the sAHP lasting for more than 1 h of drug washout. Preincubation with the specific mGluR5 antagonist, 2-methyl-6-(phenylethynyl)-pyridine (MPEP), reduced but did not completely prevent the effects of DHPG. However, preincubation with both MPEP and the mGluR1 antagonist LY367385 completely prevented the DHPG-induced changes. These results demonstrate that the DHPG-induced changes are mediated partly by mGluR5 and partly by mGluR1. Because Group I mGluRs are linked to PLC via G-protein activation, we also investigated pathways downstream of PLC activation, using chelerythrine and cyclopiazonic acid to block protein kinase C (PKC) and inositol 1,4,5-trisphosphate-(IP 3 )-activated Ca 2+ stores, respectively. Neither inhibitor affected the DHPG-induced suppression of the sAHP or the increase in excitability nor did an inhibitor of PLC itself, U-73122. Taken together, these results argue that in CA1 pyramidal cells in the adult rat, DHPG activates mGluRs of both the mGluR5 and mGluR1 subtypes, causing a long-lasting suppression of the sAHP and a consequent persistent increase in excitability via a PLC-, PKC-, and IP 3 -independent transduction pathway. Footnotes Address for reprint requests: D. R. Ireland, Dept. Psychology, University of Otago, P.O. Box 56, Dunedin, New Zealand (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Nelson, Thomas E.; Ur, Christina L.; Gruol, Donna L.

doi: N/Apmid: 12091569

Abstract The cytokine interleukin-6 (IL-6) is chronically expressed at elevated levels within the CNS in many neurological disorders and may contribute to the histopathological, pathophysiological, and cognitive deficits associated with such disorders. However, the effects of chronic IL-6 exposure on neuronal function in the CNS are largely unknown. Therefore using intracellular recording and calcium imaging techniques, we investigated the effects of chronic IL-6 exposure on the physiological properties of cerebellar Purkinje neurons in primary culture. Two weeks of exposure to 1,000 units/ml (U/ml) IL-6 resulted in altered electrophysiological properties of Purkinje neurons, including a significant reduction in action potential generation, an increase in input resistance, and an enhanced electrical response to the ionotropic glutamate receptor agonist, α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) compared with untreated neurons. Lower concentrations of IL-6 (100 and 500 U/ml) had no effects on these electrophysiological parameters. However, neurons exposed to 500 U/ml chronic IL-6 resulted in significantly elevated resting levels of intracellular calcium as well as an increase in the intracellular calcium signal of Purkinje neurons in response to AMPA, effects not observed in neurons exposed to 1,000 U/ml chronic IL-6. Morphometric analysis revealed a lack of gross structural changes following chronic IL-6 treatment, such as in the number, size, and extent of dendritic arborization of Purkinje neurons in culture. Using immunohistochemistry, we found that cultured Purkinje neurons express both the IL-6 receptor and its intracellular signaling subunit, gp130, indicating that IL-6 may act directly on Purkinje neurons to alter their physiological properties. The present data show that chronic exposure to elevated levels of IL-6, such as occurs in various neurological diseases, produces alterations in several important physiological properties of Purkinje neurons and that these changes occur in the absence of neuronal toxicity, damage, or death. The results support the hypothesis that chronic IL-6 exposure can disrupt normal CNS function and thereby contribute to the pathophysiology associated with many neurological diseases. Footnotes Address for reprint requests: D. L. Gruol, Dept. of Neuropharmacology, CVN-11, The Scripps Research Institute, 10550 North Torrey Pines Rd., La Jolla, CA 92037 (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Stemkowski, Patrick L.; Tse, Frederick W.; Peuckmann, Vera; Ford, Christopher P.; Colmers, William F.; Smith, Peter A.

doi: N/Apmid: 12091553

Abstract Suppression of the voltage-activated, noninactivating K + conductance (M conductance; g M ) by muscarinic agonists, P 2Y agonists or bradykinin increases neuronal excitability. All agonist effects are mediated, at least in part, via the Gq/ 11 class of G protein. We found, using whole cell or perforated patch recording from bullfrog sympathetic B neurons that ATP-induced suppression of g M was attenuated by the phospholipase C (PLC) inhibitor, U73122 (IC 50 ∼0.14 μM) but not by the inactive isomer, U73343 . The ability of extracellularly applied U73122 to inhibit PLC was confirmed by its antagonism of ATP-induced elevation of intracellular Ca 2+ as measured by fura-2 photometry. ATP-induced g M suppression was not antagonized by the protein kinase C (PKC) inhibitor, chelerythrine (5 μM extracellular +10 μM intracellular), by the Ca 2+ -ATPase inhibitor, thapsigargin (5 μM), or by inositol trisphosphate (InsP 3 ) receptor antagonists, heparin (∼300 μM) or xestospongin C (1.8 μM). The effect of ATP on g M was thus dependent on PLC yet independent of PKC and of InsP 3 -induced release of intracellular Ca 2+ . We therefore tested the involvement of a PKC-independent action of diacylglycerol (DAG) that could occur via activation of Ras. This low-molecular-weight G protein is activated following DAG binding to Ras -GRP, a neuronal Ras -GTP exchange factor. However, impairment of Ras function by culturing neurons with isoprenylation inhibitors (perillic acid, 0.1 mM, or α-hydroxyfarnesyl-phosphonic acid, 10 μM) failed to affect ATP-induced g M suppression. Inhibition of MEK (mitogen-activated protein kinase), a downstream target of Ras , by using PD 98059 (10 μM) was also ineffective. The transduction mechanism used by ATP to suppress g M in frog sympathetic neurons therefore differs from the PLC-independent mechanism used by muscarine and from the PLC and Ca 2+ -dependent mechanism used by bradykinin and UTP in mammalian ganglia. The possibility remains that “lipid-signaling” mechanisms, perhaps involving PLC-induced depletion of phosphatidylinositol bisphosphate, are involved in PLC-mediated inhibition of g M by ATP in amphibian sympathetic neurons. Footnotes Address for reprint requests: P. A. Smith, Dept. of Pharmacology, University of Alberta, 9.75 Medical Sciences Bldg., Edmonton, Alberta T6G 2H7, Canada (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Chen, Jen-I; Ha, Brian; Bushnell, M. Catherine; Pike, Bruce; Duncan, Gary H.

doi: N/Apmid: 12091568

Abstract The role of the somatosensory cortices (SI and SII) in pain perception has long been in dispute. Human imaging studies demonstrate activation of SI and SII associated with painful stimuli, but results have been variable, and the functional relevance of any such activation is uncertain. The present study addresses this issue by testing whether the time course of somatosensory activation, evoked by painful heat and nonpainful tactile stimuli, is sufficient to discriminate temporal differences that characterize the perception of these stimulus modalities. Four normal subjects each participated in three functional magnetic resonance imaging (fMRI) sessions, in which painful (noxious heat 45–46°C) and nonpainful test stimuli (brushing at 2 Hz) were applied repeatedly (9-s stimulus duration) to the left leg in separate experiments. Activation maps were generated comparing painful to neutral heat (35°C) and nonpainful brushing to rest. Directed searches were performed in SI and SII for sites reliably activated by noxious heat and brush stimuli, and stimulus-dependent regions of interest (ROI) were then constructed for each subject. The time course, per stimulus cycle, was extracted from these ROIs and compared across subjects, stimulus modalities, and cortical regions. Both innocuous brushing and noxious heat produced significant activation within contralateral SI and SII. The time course of brush-evoked responses revealed a consistent single peak of activity, approximately 10 s after the onset of the stimulus, which rapidly diminished upon stimulus withdrawal. In contrast, the response to heat pain in both SI and SII was characterized by a double-peaked time course in which the maximum response (the 2nd peak) was consistently observed ∼17 s after the onset of the stimulus (8 s following termination of the stimulus). This prolonged period of activation paralleled the perception of increasing pain intensity that persists even after stimulus offset. On the other hand, the temporal profile of the initial minor peak in pain-related activation closely matched that of the brush-evoked activity, suggesting a possible relationship to tactile components of the thermal stimulation procedure. These data indicate that both SI and SII cortices are involved in the processing of nociceptive information and are consistent with a role for these structures in the perception of temporal aspects of pain intensity. Footnotes Address for reprint requests: G. H. Duncan, Centre de recherche en sciences neurologiques, C.P. 6128, Succursale Centre-Ville, Université de Montréal, Montréal, Québec H3C 3J7 Canada (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Crowder, Tara L.; Weiner, Jeff L.

doi: N/Apmid: 12091531

Abstract The nucleus accumbens, a brain region involved in motivation, attention, and reward, receives substantial glutamatergic innervation from many limbic structures. This excitatory glutamatergic input plays an integral role in both normal and pathophysiological states. Despite the importance of glutamatergic transmission in the nucleus accumbens, the specific receptor subtypes that mediate glutamatergic signaling in this brain region have not been fully characterized. The current study sought to examine the possible role of the kainate subclass of glutamate receptor in the nucleus accumbens. Kainate receptors are relatively poorly understood members of the ionotropic glutamate receptor family and are highly expressed in the nucleus accumbens. Recent studies have highlighted a number of novel pre- and postsynaptic functions of kainate receptors in several other brain regions. Using the whole cell patch-clamp technique, we report the first demonstration of functional kainate receptors on neurons within the core region of the nucleus accumbens. In addition, we present evidence that activation of kainate receptors in this brain region inhibits excitatory synaptic transmission via a presynaptic mechanism. Footnotes Address for reprint requests: J. L. Weiner, Dept. of Physiology and Pharmacology, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1083 (E-mail: [email protected] ). Copyright © 2002 The American Physiological Society

Hornby, T. George; McDonagh, Jennifer C.; Reinking, Robert M.; Stuart, Douglas G.

doi: N/Apmid: 12091534

Abstract The purpose of this study was to quantify the effects of excitatory modulation on the intrinsic properties of motoneurons (MNs) in slices of spinal cord taken from the adult turtle. Responses were noted following application of an excitatory modulator: serotonin (5-HT), muscarine, trans -1-amino-1,3-cyclopentane dicarboxylic acid (tACPD), or all three combined. A sample of 44 MNs was divided into 2 groups, on the basis of whether MNs did (28/44) or did not (16/44) demonstrate a nifedipine-sensitive acceleration of discharge during a 2-s, intracellularly injected stimulus pulse. Such acceleration indicates the development of a plateau potential (PP). Excitatory modulation lowered the MNs' resting potential, increased input resistance, decreased rheobase, reduced several afterhyperpolarization values, and shifted the conventional, one-phase stimulus current–spike frequency ( I-f ) relation to the left. For both MN groups, the relative efficacy of excitatory modulation on both non-PP and PP MNs was generally in the following order: combined application > 5-HT ≈ muscarine > tACPD. In many instances, the effects of modulation differed significantly for non-PP versus PP MNs, the most pronounced being in their I-f relation. To describe this difference, it was necessary to measure a two-phase relation. In PP MNs, excitatory modulation considerably increased the slope of the first (initial) phase and flattened the second (later) phase of this relation. The latter result bore similarities to that obtained in a previous study, which addressed MN firing behavior during fictive locomotion of the high-decerebrate cat. Footnotes Address for reprint requests: D. G. Stuart, Dept. of Physiology, The Univ. of Arizona College of Medicine, Tucson, AZ 85724-5051 (E-mail: [email protected] ). Present addresses: T. G. Hornby, Dept. of Physical Medicine and Rehabilitation, Northwestern University, 345 East Superior, Chicago, IL 60611; J. C. McDonagh, Arizona School of Health Sciences, 5850 E. Still Circle, Mesa, AZ 85206; R. M. Reinking, Program in Applied Mathematics, The University of Arizona, Tucson, AZ 85721-0089. Copyright © 2002 The American Physiological Society