Khodakhah Lab and the Cerebellum: A Love Story

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Khodakhah Lab and the Cerebellum A Love Story

• AECOM, Kennedy 506. Beautiful view of parking
  lot. http//www.aecom.yu.edu/kamlab
• Karina Alviña, Paola Calderón, Johanna Dizon,
  Sung-Min Park, Esra Tara, Joy Walter, and Kamran
  Khodakhah

Typical PC response to a single pulse electrical
GC layer stimulation
stimulus
90 µA stimulation
Anatomy of the cerebellum
200 µV
50 ms
Average PC Response vs. Normalized stimulus
intensity
Raster plot of PC responses to different GC
stimulation intensities
- each line represents an action potential -
stimulation occurs at time point 0
Mechanisms underlying cerebellar ataxia
Examining inhibitions role in Purkinje
cell firing rate modulation The cerebellum
coordinates the body through its principal
neuron, the Purkinje cell. The Purkinje cell
integrates over 150,000 cortical and sensory
inputs to generate the signals for coordinated
movement. Those signals are in the form of a
rate code that modulates from the intrinsic rate
of 50 Hz to span from 1 to 250 Hz. Firing
rates under 50 Hz require some form of net
inhibition from inhibitory interneurons basket
cells and stellate cells. However, their
strengths relative to each other and the role of
a single interneuron in Purkinje cell firing rate
modulation remain unclear. We will clarify these
issues using a spike train stimulus protocol in
basket and stellate cells while recording from a
connected Purkinje cell. Question Can a single
interneuron sustain a GABA conductance to shunt a
Purkinje cells intrinsic inward currents? A

B
Cerebellar ataxia is a disorder characterized by
poorly coordinated movements and impairment of
balance and gait. Mutations in the P/Q-type
calcium channel cause ataxia in both humans and
mice. These mutations result in decreased calcium
currents in Purkinje cells of ataxic animals.
However, the mechanisms by which these
alterations produce ataxia are largely unknown.
Previous work in our lab has shown that
P/Q-type calcium channels are required to sustain
the normal intrinsic activity of cerebellar
Purkinje cells. For that reason, we are
evaluating the hypothesis that disruptions in
Purkinje cells normal firing result in the motor
impairment observed in P/Q-type calcium channel
ataxic mutants. We monitored the spontaneous
activity of mutant Purkinje cells using
extracellular recording in cerebellar slices
(Figure 1). Compared to normal littermates,
mutants showed an increased variation between
action potentials. In ducky mutants, the
observed reduction in the density of calcium
current could result in decreased activation of
the calcium-activated potassium channels that set
the interspike interval. Therefore, we decided to
test whether the activation of them could recover
the regular firing rate in mutant Purkinje cells
(Figure 2). In addition, we perfused chronically
the cerebellum of ducky heterozygous and tested
the motor performance using two common paradigms
to assess cerebellar-mediated motor coordination,
accelerating rotarod and balance beam (Figure 3).
Mapping granule cell-to-Purkinje cell connectivity
We are characterizing the functional
connectivity between GCs and PCs, thereby
addressing three controversial questions 1) Does
activity in this pathway propagate in a dispersed
or patchy manner? 2) Do the ascending axon and
parallel fiber regions of the GC provide
differential input to PCs? and 3) How do
molecular layer inhibitory interneurons between
GCs and PCs modulate PC output? In ongoing
experiments, PC electrical activity in response
to stimulation in multiple underlying GC patches
is monitored via extracellular recording.

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2b
Figure 2. Plots of instantaneous firing rate vs.
time after stimulus onset corresponding to the 63
stimulation sites are overlayed on a coordinate
map of the PC and the surrounding layers. These
data were obtained from the same cell, in the
absence (2a) or presence (2b) of picrotoxin, a
GABAA receptor blocker. Blue and red traces
denote PC excitation and inhibition,
respectively.
Figure 1. Caged glutamate is released by a 1
ms-40 µm diameter UV pulse at 63 sites on a 160 x
200 µm granule cell region underlying the
Purkinje cell being assessed.
Mechanisms underlying episodic neurological
disorders
A. Recording configuration Purkinje cells are
voltage-clamped at -60 mV with a high Cl internal
solution. Interneurons are stimulated either
extracellularly or in a loose-patch
configuration. B. PC IPSCs elicited from an
interneuron (basket cell) stimulated at 100 Hz
for 1 second (only first four IPSCs shown). Note
that the IPSCs can be evoked before the prior
IPSC fully decaysthereby maintaining a
continuous GABAA shunt (baseline shown as dotted
line).
Episodic neurological disorders such as
episodic ataxia and paroxysmal dyskinesia are
characterized by transient expression of
symptoms superimposed on a normal baseline.
The episodic nature of these disorders is due
to a transient dysfunction of ion channels.
While the symptoms of this family of disorders
are different, they share common triggers such
as psychological stress, caffeine and alcohol.
We propose to investigate the mechanism by
which these different triggers lead to the
expression of identical symptoms. The ataxic
mouse tottering inherits a mutation in P/Q type
calcium channels, which results in a reduction
in the P/Q channel current density in cerebellar
Purkinje cells and causes an ataxic baseline
punctuated by episodes of severe dyskinesia.
We hypothesize that chemical, physical and
psychological stressors trigger episodes of
dyskinesia via a common physiological pathway,
by altering the activity of Purkinje cells. To
address this question, we use the tottering mouse
as a model of episodic neurological disorders
and record single unit Purkinje cell activity in
vivo in awake behaving tottering mice before,
during and following an attack triggered by
caffeine, ethanol or stress.
Peaks and Valleys of the GABA shunt at different
stimulation frequencies
Filled data points represent the peaks of each
IPSCthat is the maximum amount of GABA-mediated
current. Open data points represent the trough
between two IPSCsthat is the minimum amount of
GABA- mediated current. Data is an average of
10 trials. Error bars are equal to SEM. Dashed
line at y 0 indicates baseline. Stimulation
frequencies were at 10, 50, and 67 Hz for 1
second. However data only to 250 ms is shown.
Figure 2. Purkinje cell activity in awake
behaving mice
Figure 1. In vivo recording of single unit
activity in awake behaving mice
A single basket cell may maintain a continuous
GABA shunt over baseline for nearly 250 ms.