Elsevier

Neuroscience

Volume 110, Issue 1, 1 March 2002, Pages 105-121
Neuroscience

Cerebellar input to magnocellular neurons in the red nucleus of the mouse: synaptic analysis in horizontal brain slices incorporating cerebello-rubral pathways

https://doi.org/10.1016/S0306-4522(01)00544-9Get rights and content

Abstract

We studied the synaptic input from the nucleus interpositus of the cerebellum to the magnocellular division of the red nucleus (RNm) in the mouse using combined electrophysiological and neuroanatomical methods. Whole-cell patch-clamp recordings were made from brain slices (125–150 μm) cut in a horizontal plane oriented to pass through both red nucleus and nucleus interpositus. Large cells that were visually selected and patched were injected with Lucifer Yellow and identified as RNm neurons.

Using anterograde tracing from nucleus interpositus in vitro, we examined the course of interposito-rubral axons which are dispersed in the superior cerebellar peduncle. In vitro monosynaptic responses in RNm were elicited by an electrode array placed contralaterally in this pathway but near the midline. Mixed excitatory post-synaptic potentials (EPSPs)/inhibitory post-synaptic potentials (IPSPs) were observed in 48 RNm neurons. Excitatory components of the evoked potentials were studied after blocking inhibitory components with picrotoxin (100 μM) and strychnine (5 μM). All RNm neurons examined continued to show monosynaptic EPSPs after non-N-methyl-D-aspartate (NMDA) glutamate receptor components were blocked with 10 μM 6,7-dinitroquinoxaline-2,3-dione or 5 μM 2,3-dihydro-6-nitro-7-sulfamoyl-benzo(f)-quinoxaline (NBQX; n=12). The residual potentials were identified as NMDA receptor components since they (i) were blocked by the addition of the NMDA receptor antagonist, D,L-2-amino-5-phosphonovaleric acid (APV), (ii) were voltage-dependent, and (iii) were enhanced by Mg2+ removal.

Inhibitory components of the evoked potentials were studied after blocking excitatory components with NBQX and APV. Under these conditions, all RNm neurons studied continued to show IPSPs. Blockade of GABAA receptors reduced but did not eliminate the IPSPs. These were eliminated when GABAA receptor blockade was combined with strychnine to eliminate glycine components of the IPSPs. Thus, IPSPs evoked by midline stimulation of the superior cerebellar peduncle, while blocking α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) and NMDA receptors, raise the possibility of direct inhibitory inputs to RNm from the cerebellum.

In summary we propose that the special properties of the NMDA receptor components are considered important for the generation of RNm motor commands: their slow time course will contribute a steady driving force for sustained discharge and their voltage dependency will facilitate abrupt transitions from a resting state of quiescence to an active state of intense motor command generation.

Section snippets

Animals

Fifty-eight C57BL/6 mice were used in the study. Pregnant females were obtained from Charles Rivers Laboratories. Pups 12–18 days old were selected for use in the slice experiments, and six of their 15–20 day old littermates were used in anterograde tracing experiments. Animals were housed in a vivarium maintained and supervised by the Center for Comparative Medicine at Northwestern University Medical Center. All experimental procedures were reviewed and approved by the University animal

Results

Figure 1 depicts the basic arrangement of the horizontal slice used for recording from RNm neurons. Usually, the landmarks that were most prominent in the living section were the third ventricle, the fasciculus retroflexus, and the indentation of the brainstem at the lateral margins of the mesencephalon that is evident just opposite the hippocampal formation. The brainstem indentation is located at a level just rostral to the decussation of the superior cerebellar peduncle and just caudal to

Discussion

In this report, a horizontal brain slice preparation was used to characterize the synaptic projection from the cerebellum to the RNm in P12–P18 mouse pups. We provide the first definitive evidence that the projection from the cerebellum to the RNm utilizes NMDA, in addition to non-NMDA receptors. There may also be a direct inhibitory projection. The combination of physiological recording and intracellular labeling provided direct anatomical verification of the identity of recorded neurons. RNm

Summary

We studied synaptic mechanisms in identified RNm neurons in response to contralateral stimulation in a horizontal brain slice that preserves axons from the cerebellar nucleus interpositus. We demonstrated that synaptic transmission in this pathway evokes NMDA, in addition to AMPA, receptor-mediated monosynaptic EPSPs and appears also to evoke monosynaptic GABAA and glycine receptor-mediated IPSPs. The EPSPs will contribute to the generation of the motor commands that emanate from this nucleus,

Acknowledgements

This work was supported by R01 NS36004, NS-HL60097 and NS-HL60969. We wish to acknowledge the invaluable contributions of Dr. S.R. Glaum (deceased) in establishing Dr. Houk’s cellular neurophysiology laboratory in which these experiments were performed. We also acknowledge the helpful advice on the horizontal slice we received from Edgar Garcia-Rill.

References (80)

  • J.C Houk

    Red nucleus: role in motor control

    Curr. Opin. Neurobiol.

    (1991)
  • J.C Houk et al.

    Distributed motor commands in the limb promotor network

    Trends Neurosci.

    (1993)
  • K Itoh et al.

    Application of coupled oxidation reaction to electron microscopic demonstration of horseradish peroxidase: Cobalt-glucose oxidase method

    Brain Res.

    (1979)
  • T Katafuchi et al.

    Responses of rat lateral hypothalamic neuron activity to vestibular nuclei stimulation

    Brain Res.

    (1987)
  • J Keifer et al.

    Intrinsic and synaptic properties of turtle red nucleus neurons in vitro

    Brain Res.

    (1993)
  • J.S King et al.

    An experimental light and electron microscopic study of cerebellorubral projections in the opossum, Didelphis marsupialis virginiana

    Brain Res.

    (1973)
  • M Kubota et al.

    Release of endogenous GABA from the cat red nucleus slices

    Brain Res.

    (1983)
  • M Kubota et al.

    Release of endogenous GABA from the cat red nucleus slices

    Brain Res.

    (1983)
  • M Kubota et al.

    Calcium-dependent potentials in mammalian red nucleus neurons in vitro

    Neurosci. Res.

    (1984)
  • K Kumoi et al.

    Immunohistochemical localization of gamma-aminobutyric acid- and aspartate-containing neurons in the rat deep cerebellar nuclei

    Brain Res.

    (1988)
  • P Mason et al.

    Intracellular responses of raphe magnus neurons during the jaw-opening reflex evoked by tooth pulp stimulation

    Brain Res.

    (1986)
  • F Murakami et al.

    Immunocytochemical demonstration of GABAergic synapses on identified rubrospinal neurons

    Brain Res.

    (1983)
  • K Sato et al.

    Localization of glycine receptor alpha 1 subunit mRNA-containing neurons in the rat brain: an analysis using in situ hybridization histochemistry

    Neuroscience

    (1991)
  • Y Shinoda et al.

    Morphology of single neurons in the cerebello-rubrospinal system

    Behav. Brain Res.

    (1988)
  • W.J Song et al.

    Segregation of cerebrorubral and cerebellorubral synaptic inputs on rubrospinal neurons of fetal cats as demonstrated by intracellular recording

    Neurosci. Lett.

    (1993)
  • J Taguchi et al.

    Immunohistochemical studies on distribution of GABAA receptor complex in the rat brain using antibody against purified GABAA receptor complex

    Brain Res.

    (1989)
  • T.M Teune et al.

    Cerebellar projections to the red nucleus and inferior olive originate from separate populations of neurons in the rat: a non-fluorescent double labeling study

    Brain Res.

    (1995)
  • N Tsukahara et al.

    Properties of cerebello–precerebellar reverberating circuits

    Brain Res.

    (1983)
  • F Viana et al.

    Postnatal changes in rat hypoglossal motoneuron membrane properties

    Neuroscience

    (1994)
  • G Vuillon-Cacciuttolo et al.

    GABA neurons in the cat red nucleus: a biochemical and immunohistochemical demonstration

    Neurosci. Lett.

    (1984)
  • C.J Wilson et al.

    Dendritic and somatic appendages of identified rubrospinal neurons of the cat

    Neuroscience

    (1987)
  • P Angaut et al.

    The dentatorubral projection. An autoradiographic study in rats

    Brain Behav. Evol.

    (1987)
  • A.L Babalian et al.

    Electrophysiologic properties of red nucleus neurons in the rat brain slices

    Neirofiziologiia

    (1991)
  • C Batini et al.

    Cerebellar nuclei and the nucleocortical projections in the rat: retrograde tracing coupled to GABA and glutamate immunohistochemistry

    J. Comp. Neurol.

    (1992)
  • J Bäurle et al.

    Differential number of glycine- and GABA-immunoreactive neurons and terminals in the deep cerebellar nuclei of normal and Purkinje cell degeneration mutant mice

    J. Comp. Neurol.

    (1997)
  • A.S Berrebi et al.

    Glycinergic innervation of red nucleus

    Soc. Neurosci. Abstr.

    (1994)
  • S Chen et al.

    Colocalization of neurotransmitters in the deep cerebellar nuclei

    J. Neurocytol.

    (1993)
  • J Davies et al.

    Amino acid receptor mediated excitatory synaptic transmission in the cat red nucleus

    J. Physiol. Lond.

    (1986)
  • R.A Deisz et al.

    Voltage dependency of excitatory postsynaptic potentials of rat neocortical neurons

    J. Neurophysiol.

    (1991)
  • Eisenman, L.N., Keifer, J., Houk, J.C., 1991. Positive feedback in the cerebro-cerebellar recurrent network may explain...
  • Cited by (23)

    • The mammalian red nucleus and its role in motor systems, including the emergence of bipedalism and language

      2012, Progress in Neurobiology
      Citation Excerpt :

      Mannen (1960) was able to use the term “closed” based on the fact that extranuclear dendrites of the cells inside the nucleus were rarely found to exist outside the nuclear boundary – that is, in the neighboring reticular formation. The mNr of other mammalian species also showed similar features of a closed nucleus (see monkey in Burman et al., 2000; mouse in Jiang et al., 2002). In Fig. 7 taken from human mNr sections, we can readily identify the same three groups first proposed by Mitomo (1942a) i.e. the dorsal, ventral and cell-poor intermediate groups.

    • Electrophysiological actions of the rubrospinal tract in the anaesthetised rat

      2008, Experimental Neurology
      Citation Excerpt :

      In the rat, they decussate at about 1 mm caudal to the RN and then run rostrally through the nucleus en route to the thalamus. The terminals on RN neurones are believed to be collaterals of thalamic afferents (Caughell and Flumerfelt, 1977; also see Shinoda et al., 1988 in the cat; Jiang et al., 2002 in the mouse). Thus, stimuli anywhere in the RN, including the parvocellular region and in more anterior positions, should be able to activate the interpositorubral fibres to the RNm neurones.

    View all citing articles on Scopus
    View full text