In vivo analysis of cerebellar circuitry has focused almost exclusively on Purkinje cells, identified by their iconic patterns of complex and simple spikes (CSs and SSs). However, views of cerebellar function based exclusively on the physiology of Purkinje cells ignore the role of interneurons and distort the attributes of cerebellar afferent systems. It is universally assumed that SSs are modulated by the activity of the mossy fiber-granule cell-parallel fiber projection to Purkinje cell dendrites. In fact, during natural vestibular stimulation, vestibular primary afferent mossy fiber afferents discharge out of phase with the SSs recorded from nodular Purkinje cells. Consequently, it is unlikely that the cerebellar output signal merely reflects a gain-controlled version of the mossy fiber input signal. We proposed that SS modulation reflects the action of climbing fibers on cerebellar interneurons. We will test specific versions of this hypothesis by recording from identified interneurons. We have three objectives. First, we will record extracellularly from interneurons in the uvula-nodulus of anesthetized mice during natural vestibular stimulation. Interneurons will be labeled juxtacellularly with neurobiotin. The depth and phase of modulation of interneuronal discharge relative to that of Purkinje cell CSs and SSs will indicate which interneurons could modulate SSs. Second, we will study how the modulated activity of interneurons and Purkinje cells is altered by a unilateral labyrinthectomy (UL). Following a UL, the ipsilateral uvula-nodulus is accessible to vestibular information mediated only by climbing fibers whose modulation depends on the contralateral, intact labyrinth. Third, we will also make microlesions in the p-nucleus and dorsomedial cell column (dmcc) in the contralateral inferior olive. This will leave one side of the cerebellum accessible to vestibular information mediated only by vestibular mossy fibers. We will compare the effects of reduced vestibular signaling on interneurons and Purkinje cells. Fourth, we will microinject miRNAs in viral vectors with a cell specific promoter to selectively reduce expression of GABA-A alpha 1 receptors in nodular Purkinje cells. Fifth, we will also use miRNAs to selectively reduce synthesis of GABA in Golgi cells. We will analyze the effects of """"""""knocking down"""""""" GABAergic signaling in these two cell types. We will characterize the stimulus-modulated functions of identified interneurons for the first time. We will interfere with cerebellar circuitry at a cellular level and test the role of interneurons in the modulation of SSs. The proposed research will speed application of molecular techniques to the treatment of patients with cerebellar disorders.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY018561-05
Application #
8113968
Study Section
Sensorimotor Integration Study Section (SMI)
Program Officer
Araj, Houmam H
Project Start
2007-08-01
Project End
2013-07-31
Budget Start
2011-08-01
Budget End
2013-07-31
Support Year
5
Fiscal Year
2011
Total Cost
$292,723
Indirect Cost
Name
Oregon Health and Science University
Department
Physiology
Type
Schools of Medicine
DUNS #
096997515
City
Portland
State
OR
Country
United States
Zip Code
97239
Barmack, N H; Yakhnitsa, V (2015) Climbing fibers mediate vestibular modulation of both ""complex"" and ""simple spikes"" in Purkinje cells. Cerebellum 14:597-612
Barmack, N H; Yakhnitsa, V (2013) Modulated discharge of Purkinje and stellate cells persists after unilateral loss of vestibular primary afferent mossy fibers in mice. J Neurophysiol 110:2257-74
Qian, Z; Micorescu, M; Yakhnitsa, V et al. (2012) Climbing fiber activity reduces 14-3-3-? regulated GABA(A) receptor phosphorylation in cerebellar Purkinje cells. Neuroscience 201:34-45
Qian, Zuyuan; Yakhnitsa, Vadim; Barmack, Neal H (2011) Climbing fiber-evoked Purkinje cell discharge reduces expression of GABA(A) receptor-associated protein and decreases its interaction with GABA(A) receptors. J Neurochem 117:197-208
Barmack, Neal H; Yakhnitsa, Vadim (2011) Microlesions of the inferior olive reduce vestibular modulation of Purkinje cell complex and simple spikes in mouse cerebellum. J Neurosci 31:9824-35
Barmack, Neal H; Yakhnitsa, Vadim (2011) Topsy turvy: functions of climbing and mossy fibers in the vestibulo-cerebellum. Neuroscientist 17:221-36
Barmack, N H; Qian, Z; Yakhnitsa, V (2010) Climbing fibers induce microRNA transcription in cerebellar Purkinje cells. Neuroscience 171:655-65
Qian, Zuyuan; Bilderback, Timothy R; Barmack, Neal H (2008) Acyl coenzyme A-binding protein (ACBP) is phosphorylated and secreted by retinal Muller astrocytes following protein kinase C activation. J Neurochem 105:1287-99
Barmack, N H; Yakhnitsa, V (2008) Distribution of granule cells projecting to focal Purkinje cells in mouse uvula-nodulus. Neuroscience 156:216-21
Barmack, Neal H; Yakhnitsa, Vadim (2008) Functions of interneurons in mouse cerebellum. J Neurosci 28:1140-52