preclinical evaluation of lobeline for the treatment of adhd

193 Pages·2016·0.76 MB·English

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Theses and Dissertations--Pharmacy College of Pharmacy
2011
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Yolanda D. Williams
University of Kentucky, [email protected]
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Williams, Yolanda D., "PRECLINICAL EVALUATION OF LOBELINE FOR THE TREATMENT OF ADHD:
COMPARISON WITH PSYCHOSTIMULANT THERAPIES" (2011). Theses and Dissertations--Pharmacy. 3.
https://uknowledge.uky.edu/pharmacy_etds/3
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The document mentioned above has been reviewed and accepted by the student’s advisor, on
behalf of the advisory committee, and by the Director of Graduate Studies (DGS), on behalf of
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Yolanda D. Williams, Student
Dr. Linda P. Dwoskin, Major Professor
Jim Pauly, Ph.D., Director of Graduate Studies
PRECLINICAL EVALUATION OF LOBELINE FOR THE TREATMENT OF
ADHD: COMPARISON WITH PSYCHOSTIMULANT THERAPIES

_________________________________________

DISSERTATION
_________________________________________

A dissertation submitted in partial fulfillment of the
requirements for the degree of Doctor of Philosophy in the
College of Pharmacy
at the University of Kentucky

By
Yolanda D. Williams

Lexington, Kentucky

Director: Dr. Linda P. Dwoskin, Professor of Pharmaceutical Sciences

Lexington, Kentucky

2011

Copyright © Yolanda D. Williams 2011
ABSTRACT OF DISSERTATION

PRECLINICAL EVALUATION OF LOBELINE FOR THE TREATMENT OF
ADHD: COMPARISON WITH PSYCHOSTIMULANT THERAPIES

This dissertation work investigated the effect of acute and repeated in vivo
administration of lobeline on dopamine transporter (DAT) and vesicular
monoamine transporter (VMAT2) function. The effects of lobeline were then
compared to the effects of acute and repeated in vivo administration of
methylphenidate and amphetamine to determine if lobeline produced similar
effects compared to these Attention Deficit Hyperactivity Disorder (ADHD)
medications. These medications are considered the first line of pharmacotherapy
for ADHD, although there is a growing concern associated with their potential for
abuse and other side effects. This merits the need for novel ADHD treatments
that have a safer side effect profile. If lobeline alters DAT and VMAT2 function in
the same way as methylphenidate or amphetamine, further investigation may be
necessary to evaluate lobeline as a potential treatment for ADHD. Kinetic
analysis of [3H]dopamine (DA) was utilized to determine the effect on DAT and
VMAT2 function in rat striatum. Results from the DAT experiments, revealed that
lobeline as well as amphetamine had no effect on DAT function. However,
methylphenidate increased DAT function after acute and 7-day treatment. None
of the drug treatment regimens altered K . To determine if the methylphenidate-
m
induced increase in DAT function was due to DAT trafficking, biotinylation and
Western blot analyses were performed. Acute administration of methylphenidate
did not alter surface DAT, however repeated administration of methylphenidate
for 7 days decreased intracellular DAT, suggesting that methylphenidate
redistributes DAT in a time-dependent manner. Similar results were found in the
VMAT2 experiments. Lobeline and amphetamine had no effect on VMAT2
function after acute or repeated administration. Amphetamine decreased the K
m
after repeated administration for 7 days. Methylphenidate increased VMAT2
function after acute and repeated administration for 7 days. The overall results of
these experiments suggest that methylphenidate interacts with DAT and VMAT2
in a different manner than amphetamine and lobeline. In addition, since lobeline
and amphetamine had no effect on DAT and VMAT2 function, further
investigation is warranted to elucidate the underlying mechanisms of the
therapeutic actions of these agents. This additional information will aid in the
development of novel treatments for ADHD.

Keywords: lobeline, methylphenidate, amphetamine, striatum, ADHD, dopamine
transporter, vesicular monoamine transporter
Yolanda D. Williams
Student’s Signature

October 25, 2011__
Date
PRECLINICAL EVALUATION OF LOBELINE FOR THE TREATMENT OF
ADHD: COMPARISON WITH PSYCHOSTIMULANT THERAPIES

By

Yolanda D. Williams

Jim Pauly, Ph.D.__________
Director of Graduate Studies

Linda P. Dwoskin, Ph.D.____
Director of Dissertation

October 25, 2011_________
ACKNOWLEDGEMENTS

Hebrews 12:1b-3
“And let us run with perseverance the race marked out for us, fixing our eyes on
Jesus, the pioneer and perfecter of faith. For the joy set before Him He endured
the cross, scorning its shame, and sat down at the right hand of God. Consider
Him who endured such opposition from sinners, so that you will not grow weary
and lose heart.”

First and foremost, I would like to thank God for continuously giving me
the strength throughout my graduate career to make it to this point. There were
numerous times in my scholastic career when I thought I could not go on, and I
believe that there is no way I would have made it without Him.

To my advisor, Dr. Linda Dwoskin, thank you for your patience and taking
the time to instill in me the qualities necessary to become a scholarly
investigator. You have taught me not just how to become an independent
researcher but to also seek guidance when necessary. You are not just my
advisor, but my friend as well. In addition, I want to express my gratitude to the
rest of my committee members: Drs. Bardo, Crooks, Martin, and Pauly for their
support and vital input throughout this journey and Dr. Craven for agreeing to be
my outside examiner.

To my family, who has been there with me every step of the way, there
are no words to express how grateful I am. To my mother, Connie Douglas and
my father, Dr. LaDell Douglas, thank you for your unconditional love and
encouragement – I love you, both. You believed in me when I didn’t believe in
myself. To my brothers, Elvan and Ethan Douglas and their families, thank you
for encouraging me to graduate by always asking me when I would be done
every time we had a conversation. To my nieces, Alyssa, Alexandra, and Alivia
and my nephews, Aidan and Julian, for giving me the motivation to finish so I can
move back home and watch you grow up instead of watching you grow up from a
distance. I want to say a very special thank you to my extended family as well
including my uncles, Dr. Costello Brown and Bernard Brown and their families for
your various ways of supporting me since I have been in graduate school. I am
truly blessed to have a family that knows the importance of higher education and
was willing to do anything in their power to help me achieve my educational goals
in life. To my cousins who have gone before me in achieving their dreams, Drs.
Eric Brown, Ninita Brown, and Linda Brown, your examples have been an
inspiration to me and I am so thankful to have you in my life.

i
To the members of Dr. Dwoskin’s lab, especially Gabi, thank you for
accommodating me with my countless experiments over the years. To the post-
docs in the lab, thank you for endless patience and advice. To my fellow
graduate students in the lab, thank you for always lending a helping hand
whenever I needed it. It was a joy to work in the lab with all of you. To the techs,
thanks for always doing anything and everything that needed to be done in the
lab. Thanks to my friends and especially my fellow classmates that started this
journey with me, Leah, Clare, Abbi, Kedra, Miranda, Brian, Stan, and Matt - I will
never forget you!

I also want to thank Catina Rossoll. She always had the answers to any
questions I had and was willing to help me in any way she could. And to my
church family, I give you a very special thank you - especially my prayer partners
for their many prayers, words of encouragement, and endless support. In
conclusion, I would like to honor the memories of my maternal and paternal
grandparents, and many others who realized the value of education when the
road was most treacherous, but paved the way for those like me so that I may
have an opportunity to achieve my dreams in life. I hope and pray that I can do
the same for those who follow behind me.

ii
TABLE OF CONTENTS

Acknowledgements............................................................................................... i

List of Tables........................................................................................................ v

List of Figures ......................................................................................................vi

I. Chapter 1: Introduction and Background
A. Attention-Deficit Hyperactivity Disorder (ADHD) ........................................1
1. Prevalence.............................................................................................1
2. Etiology..................................................................................................2
3. Symptomology and Diagnosis ..............................................................7
4. Neuroanatomical and Neurochemical Basis of ADHD ........................13
a. Neuroanatomy ...............................................................................13
b. Neurochemistry .............................................................................13
i. Dopamine synthesis and metabolism ........................................13
ii. Dopamine receptors...................................................................15
iii. Dopamine transporter ..............................................................16
iv. Norepinephrine transporter ......................................................22
v. Vesicular monoamine transporter .............................................24
5. Pharmacotherapies for ADHD .............................................................25
a. Methylphenidate ............................................................................27
i. Historical background.................................................................27
ii. Formulations..............................................................................27
iii. Pharmacokinetics and mechanism of action ............................28
b. Amphetamine ................................................................................34
i. Historical background.................................................................34
ii. Formulations .............................................................................35
iii. Pharmacokinetics and mechanism of action ............................35
6. Alternative Therapies ..........................................................................41
7. Investigational Treatments ..................................................................48
8. Lobeline as Candidate for ADHD Pharmacotherapy............................50
a. Historical background and clinical uses .........................................50
b. Pharmacokinetics and mechanism of action .................................51
9. Animal Models for ADHD ....................................................................55
a. Neonatal 6-OHDA ..........................................................................56
b. Neonatal alternative models ..........................................................57
c. Genetic models ..............................................................................58
10. Specific Aims and Hypotheses .........................................................64

II. Chapter 2: Effect of Acute and Repeated In Vivo Administration of Lobeline,
Methylphenidate and Amphetamine on Dopamine Transporter Function
A. Introduction..............................................................................................69
B. Methods ..................................................................................................70
C. Data Analysis...........................................................................................73
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D. Results.....................................................................................................73
E. Discussion...............................................................................................75

III. Chapter 3: Effect of Acute and Repeated In Vivo Administration of
Methylphenidate on Dopamine Transporter Trafficking
A. Introduction..............................................................................................97
B. Methods ..................................................................................................97
C. Data Analysis ........................................................................................100
D. Results ..................................................................................................100
E. Discussion ............................................................................................101

IV. Chapter 4: Effect of Acute and Repeated In Vivo Administration of Lobeline,
Methylphenidate, and Amphetamine on Vesicular Monoamine Transporter
Function
A. Introduction............................................................................................106
B. Methods.................................................................................................108
C. Data Analysis.........................................................................................111
D. Results...................................................................................................111
E. Discussion ............................................................................................112

V. Chapter 5: Discussion and Conclusions
A. Summary...............................................................................................130
B. Potential Mechanism for the Methylphenidate-Induced Increase
in DAT Function ....................................................................................132
1. Methylphenidate increase in DAT function is trafficking
dependent ........................................................................................134
C. Potential Mechanism for the Methylphenidate-Induced Increase in
VMAT2 Function....................................................................................137
D. Implications............................................................................................138
E. Limitations..............................................................................................143
F. Future Directions ...................................................................................144

References.......................................................................................................147
Vita...................................................................................................................177
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