Introduction to Basics of Pharmacology and Toxicology: Volume 3: Experimental Pharmacology : Research Methodology and Biostatistics

Foreword
Preface to Volume 1: General and Molecular Pharmacology: Principles of Drug Action (https://link.springer.com/book/10.1007/978...
Preface to Volume 2
Preface
Acknowledgments
Contents
Editors and Contributors
Part I: Experimental Pharmacology
1: Experimental Methodologies Involved in the Discovery of Drugs
1.1 Basic Principles of Drug Discovery
1.2 In-silico Techniques
1.2.1 Quantitative Structure-Activity Relationship (QSAR)
1.2.2 2D-QSAR and 3D-QSAR
1.2.3 Molecular Docking
1.2.4 Virtual High Throughput Screening
1.2.5 Pharmacophore Mapping
1.3 In-vitro Techniques
1.3.1 Absorption Assays
1.3.2 Metabolic Stability Assays
1.3.3 CYP450 Pathway Elucidation
1.3.4 Inhibition Potential
1.3.5 Induction Potential
1.3.6 Metabolic Profiling
1.3.7 Antisense Technology
1.4 In-vivo Techniques
1.4.1 Gene Knockout Animals
1.4.2 Transgenic Animals
1.4.3 Monoclonal Antibodies Mediated Gene Manipulation
1.5 Assays
Bibliography
2: Laboratory Animals
2.1 Introduction
2.2 Mouse
2.2.1 Scientific Name
2.2.2 Description
2.2.3 Salient Points of the Mouse as Experimental Models
2.2.4 Outbred Stocks of the Mouse
2.2.5 Inbred Strains of the Mouse
2.2.6 Genetically Modified Mouse
2.2.7 Limitations of the Mouse as Experimental Models for Human Diseases
2.3 Rat
2.3.1 Scientific Name
2.3.2 Physical Description
2.3.3 Salient Points of the Rat as an Experimental Animal
2.3.4 Outbred Stocks of Rat
2.3.5 Inbred Strains of Rat
2.3.6 Transgenic and Knock-out Rats
2.3.7 Limitation of the Rat as an Experimental Animal
2.4 Guinea Pig
2.4.1 Scientific Name
2.4.2 Description
2.4.3 Salient Points of the Guinea Pig as an Experimental Animal
2.4.4 Strains of Guinea Pig
2.5 Hamster
2.5.1 Scientific Name
2.5.2 Description
2.5.3 Salient Points of Hamster as an Experimental Animal
2.6 Gerbil
2.7 Rabbit
2.8 Cats
2.9 Pig
2.10 Dog
2.11 Monkeys and Other Non-human Primates
2.12 Horse
2.13 Sheep
2.14 Chicken
2.15 Pigeon
2.16 Frog
2.17 Zebra Fish
Bibliography
3: Care and Handling of Laboratory Animals
3.1 Introduction
3.2 Animal Ethics
3.3 Housing and Environment
3.4 Handling of Commonly Used Animals in Biomedical Research
3.4.1 Handling of Mice
3.4.2 Handling of Rat
3.4.3 Handling of Guinea Pig
3.4.4 Handling of Rabbit
3.4.5 Handling of Hamster
3.4.6 Handling of Gerbils
3.4.7 Handling of Ferret
3.4.8 Handling of Dog
3.4.9 Handling of Cat
3.5 Animal Welfare
Bibliography
4: Biological Sample Collection from Experimental Animals
4.1 Introduction
4.2 Collection of Blood
4.2.1 General Principles
4.2.2 Blood Sample Collection Through Saphenous Vein
Caution:
4.2.3 Blood Sample Collection Through Dorsal Pedal Vein
Caution:
4.2.4 Blood Sample Collection Through Tail Vein
Caution:
4.2.5 Blood Sample Collection Using a Tail Bleed (Tail Snip and Tail Nick) Method
Caution:
4.2.6 Blood Sample Collection Through Orbital Sinus
Caution:
4.2.7 Blood Sample Collection Through Jugular Vein
Caution:
4.2.8 Blood Sample Collection Using a Temporary Cannula
4.2.9 Tarsal Vein Blood Sample Collection
Caution:
4.2.10 Marginal Ear Vein/Artery Blood Sample Collection
4.2.11 Blood Sample Collection Through Cephalic Vein
4.2.12 Blood Sample Collection Through Cranial Vena Cava
4.2.13 Blood Sample Collection Through Posterior Vena Cava
4.2.14 Blood Sample Collection Through Cardiac Puncture
4.2.15 Adverse Effects of Blood Sample Collections
4.3 Urine Sample Collecting Methods
4.4 Saliva Sample Collection Method
4.5 Conclusion
Bibliography
5: Anesthesia and Euthanasia of Experimental Animals
5.1 Anesthesia of Experimental Animals
5.1.1 Anesthesia
5.1.2 Stages of Anesthesia
5.1.3 Pre-Anesthetic Medication
5.1.4 Types of Anesthesia
5.1.4.1 Local Anesthesia
5.1.4.2 General Anesthesia
5.1.5 Monitoring of Depth of Anesthesia
5.1.6 Postoperative Analgesia
5.2 Euthanasia of Experimental Animals
Bibliography
6: Equipments in Experimental Pharmacology
6.1 Organ Bath
6.1.1 History
6.1.2 Contents
6.2 Kymograph
6.2.1 History
6.2.2 Procedure
6.3 Tissue Levers
6.3.1 Magnification
6.3.2 Levers Used in Experimental Pharmacology
6.4 Analgesiometer
6.4.1 Basic Principle
6.4.2 Hot Plate Analgesiometer
6.4.3 Procedure
6.5 Digital Analgesiometer:Tail-Flick Method
6.5.1 Procedure
6.6 Electroconvulsiometer
6.6.1 Principle
6.6.2 Procedure
6.6.3 Threshold for Electroconvulsions
6.7 Elevated Plus-Maze
6.7.1 Equipment
6.7.2 Principle
6.7.3 Procedure
6.8 Actophotometer
6.8.1 Principle
6.8.2 Procedure
6.9 Rota-Rod Apparatus
6.9.1 Equipment
6.9.2 Principle
6.9.3 Procedure
6.10 Grip Strength Meter
6.10.1 Principle
6.10.2 Procedure
6.11 Cook´s Pole-Climbing Apparatus
6.11.1 Principle
6.11.2 Equipment
6.11.3 Procedure
6.11.4 Experiment
Bibliography
7: Bioassay and Experiments on Isolated Muscle Preparations
7.1 Definition
7.2 Classification
7.3 Indications
7.4 Principles
7.5 Purpose
7.6 Methods of Bioassay for Agonist
7.6.1 Direct-End-Point Method
7.6.2 Matching Method
7.6.3 Graphical Method
7.6.4 Multiple-Point Assays
7.7 Standard Isolated Muscle Preparations
7.7.1 Terminologies
7.7.2 Muscle Preparations Used in Bioassay
Bibliography
8: Experiments on Anaesthetized Intact Animals
8.1 Introduction
8.2 Intact Animal Models n Dog
8.2.1 Blood Pressure Recording in the Dog
8.2.2 Spleen Volume Measurement
8.2.3 Diuretic Study in Dogs
8.3 Intact Animal Models in Cat
8.3.1 Nictitating Membrane
8.3.2 Spinalisation of the Cat
8.3.3 Open Chest Preparation
8.3.4 Muscle Nerve Preparations
8.3.5 Models to Study the Action of Centrally Acting Skeletal Muscle Relaxants
8.3.6 Hindquarters Perfusion Model
8.3.7 Domenjoz Method for Antitussive Action
8.4 Intact Animal Models in Guinea Pigs
8.4.1 Konzett-Rossler Method for Bronchodilator Activity
8.5 Intact Animal Models in Rats
8.5.1 Blood Pressure Recording in Rats
8.6 Conclusion
Bibliography
9: Bioassay of Standard Compounds
9.1 Bioassay on Acetylcholine
9.2 Bioassay of Adrenaline
9.3 Bioassay of Histamine
9.4 Bioassay of 5HT (Serotonin)
9.5 Angiotensin II Bioassay
9.6 Bioassay of Vasopressin
9.7 Bioassay of ACTH
9.8 Bioassay of Estrogen
9.9 Bioassay of Oxytocin
Bibliography
10: Principles of EC50, ED50, pD2 and pA2 Values of Drugs
10.1 The Concept of ``Dose Descriptors´´
10.2 Preclinical Toxicity Studies
Bibliography
11: Physiological Salt Solutions
11.1 Introduction
11.2 General Principles in Preparation of Physiological Salt Solution
11.3 Commonly Used Physiological Salt Solution
11.4 Choice of Physiological Salt Solution for Specific Tissues
Bibliography
12: Validation of Animal Models
12.1 Introduction
12.2 Animal Model
12.2.1 History -Animals in Research
12.2.2 Classification of Animal Model
12.2.3 Utilization of Animal Models in Scientific Field
12.2.4 Ideal Animal Model
12.3 Validation of Animal Model
12.3.1 Face Validity
12.3.2 Construct Validity
12.3.3 Predictive Validity
12.3.4 Internal Validity
12.3.5 Factors Affecting Internal Validity
12.4 Process of Developing an Animal Model
12.5 Validation Process for Predictive Validity
12.6 Framework to Identify Models of Disease (FIMD)
12.7 Evaluation of Valid Animal Model
12.8 Limitations of Animal Model
12.9 Conclusion
Bibliography
13: Screening Methods for the Evaluation of Antidepressant Drugs
13.1 Introduction
13.2 Animal Models of Depression
13.3 Criteria for Valid Animal Models of Depression
13.4 Genetic Models
13.4.1 Traditional Genetic Mouse Models
13.4.2 Models Using Optogenetic Tools
13.4.3 Selective Breeding
13.5 Stress Exposure
13.5.1 Models of Acute Stress
13.5.1.1 Water Wheel Model
13.5.1.2 Forced Swim Test
13.5.1.3 Tail Suspension Test
13.5.1.4 Learned Helplessness Test
13.5.2 Chronic Stress Models
13.5.2.1 Chronic Mild Stress Model (CMS)
13.5.2.2 Chronic Restraint Stress Model
13.5.2.3 Chronic Social Defeat Stress
13.5.3 Isolation Induced Hyperactivity
13.5.4 Early Life Stress Model
13.6 Secondary or Iatrogenic Depression Models
13.6.1 Hormones of HPA Axis
13.6.2 Retinoic Acid Derivatives
13.6.3 Cytokines and Immune System Dysregulation
13.7 Pharmacological Models
13.7.1 Reserpine Induced Hypothermia
13.7.2 Amphetamine Potentiation
13.7.3 Apomorphine Antagonism
13.7.4 Psychostimulant Withdrawal Paradigm
13.8 Miscellaneous
13.8.1 Resident Intruder Paradigm in Rats
13.8.2 Muricidal Behavior in Rats
13.8.3 Olfactory Bulbectomy Model
13.8.4 Disruption of Circadian Rhythm
13.9 Behavioral Endpoints
13.10 Conclusion
Bibliography
14: Screening Methods for the Evaluation of Antiepileptic Drugs
14.1 Introduction
14.2 In-vitro Methods
14.2.1 GABAA Receptor Binding Measurement in Rat Brain
14.2.2 GABAB Receptor Binding Measurement in Rat Brain
14.2.3 GABA Uptake in the Cerebral Cortex of the Rat
14.2.4 GABA Release in Hippocampal Slices of Rats
14.2.5 [3H]-CPP Binding Assay for Glutamate Receptors in Rat Cerebral ortex
14.2.6 [3H]-TCP Binding Assay for NMDA Receptor Complex in Rat Cerebral Cortex
14.2.7 [H3]-Glycine Binding Assay for NMDA Receptor Complex in Rat Cerebral Cortex
14.2.8 [H3]-Strychnine Binding Assay for the Strychnine-Sensitive-Glycine Receptor in Rat Cerebral Cortex
14.2.9 [35S]-TBPS Binding Assay for Picrotoxin Site in GABA Receptor Complex in Rat Cerebral Cortex
14.2.10 Other In Vitro Techniques
14.3 In Vivo Techniques
14.3.1 Electrically Induced Seizure Models
14.3.2 Chemically Induced Seizure Models
14.3.3 Seizures Induced by Surgery
14.3.3.1 Seizures Induced by Focal Lesions
14.3.3.2 Seizures Induced by Kindling
14.3.3.3 Post Hypoxic Myoclonus in Rats
14.4 Classification of Animal Models Based on the Type of Seizure Disorder
14.4.1 Models Used for Status Epilepticus
14.4.2 Model for Infantile Spasm
14.4.2.1 Genetic Models of Chronic Infantile Spasms:
14.4.3 Models for GTCS
14.4.4 Models for Complex Partial Seizures
14.4.5 Models for Absence Seizures
14.4.6 Models for Simple Partial Seizures
14.5 Genetic Models of Epilepsy
14.5.1 Genetic Animals Models with Spontaneous Seizures
14.5.2 Genetic Animal Models with Absence Like Seizures and Tonic Convulsions
14.5.3 Genetic Animal Models with Reflex Seizures
14.6 Conclusion
Bibliography
15: Screening Methods for the Evaluation of General Anaesthetics
15.1 Introduction
15.2 Screening of Intravenous Anaesthetic Agents
15.2.1 Evaluation of Intravenous Anaesthetic Agents in Mice
15.2.2 Electroencephalographic (EEG) Threshold Test in Rats
15.2.3 Evaluation of Safety of Intravenous Anaesthetics in Rabbits
15.3 Screening of Inhalational Anaesthetic Agents
15.3.1 Screening of Volatile Anaesthetics in Mice or Rats
15.3.2 Determination of Minimal Alveolar Anaesthetic Concentration (MAC) in Rats
15.3.3 Evaluation of Safety of Inhalational Anaesthetics in Rats
Bibliography
16: Screening Methods for the Evaluation of Antipsychotic Drugs
16.1 Introduction
16.2 In-Vitro Studies
16.2.1 D1 Receptor Assay Using [H]-SCH-23390 in Rat Brain
16.2.2 D2 Receptor Assay Using 3[H]-Spiroperidol in Rat Brain
16.2.3 D3 Receptor Assay Using 3[H]-Nafadotride in Rat Brain
16.2.4 D4 Receptor Assay Using 3[H]-L-745-870
16.2.5 In-Vitro Assay for Other Receptors Involved in the Evaluation of Antipsychotic Action
16.3 Invivo: Behavioral Tests
16.3.1 Golden Hamster Test
16.3.2 Cotton Rat Test
16.3.3 Artificial Hibernation in Rats
16.3.4 Catalepsy in Rodents
16.3.5 Pole-Climb Avoidance in Rats
16.3.6 Brain Self-Stimulation Test in Rats
16.3.7 Pre-Pulse Inhibition of Startle Response in Rats
16.3.8 Other Behavioral Tests Used
16.4 In vivo Test Based on Mechanism of Action
16.5 Genetic Models of Psychosis
16.5.1 Dominant-Negative DISC-1 Transgenic Mice
16.5.2 Gsα-Transgenic Mice
16.5.3 Sandy Mice or Dysbindin-1 Mutant (DTNBP-1) Knockout Mice
16.5.4 NR-1 Hypomorphic (NR-1H) Mice
16.5.5 Heterozygous Reeler Mouse
16.5.6 Other Genetic Animal Models of Psychosis
Bibliography
17: Screening Methods for the Evaluation of Sedative-Hypnotics
17.1 Introduction
17.2 In-vitro Studies
17.2.1 GABAA Receptor Binding Measurement in Rat Brain
17.2.2 GABAB Receptor Binding Measurement in Rat Brain
17.2.3 [3H]-Mesulergine Binding Assay for 5HT2C Receptors
17.2.4 [3H]-Mepyramine Binding Assay for H1 Receptors
17.3 In-Vivo Tests for Sedative Activity
17.3.1 Effects on Motility (Sedative or Stimulatory Activity)
17.3.1.1 Method of Intermittent Observations
17.3.1.2 Open Field Test
17.3.1.3 Hole-Board Test
17.3.1.4 Combined Open Field Test
17.3.1.5 EEG Analysis from Rat Brain by Telemetry
17.3.2 Tests for Muscle Coordination
17.3.2.1 Inclined Plane
17.3.2.2 Chimney Test
17.3.2.3 Grip Strength Test
17.3.2.4 Rotarod Method: (Refer to Chap. 42)
17.4 In-vivo Tests for Hypnotic Activity
17.4.1 General Considerations
17.4.2 Potentiation of Hexobarbital Sleeping Time
17.4.3 Experimental Insomnia in Rats
17.4.4 EEG Registration in Conscious Cats
17.4.5 Automated Rat Sleep Analysis System
Bibliography
18: Screening Methods for the Evaluation of Anxiolytic Drugs
18.1 Introduction
18.2 Light/Dark Exploration Test
18.3 Elevated Plus Maze Test
18.4 Four-Plate-Test
18.5 Fear Potentiated Startle Test
18.6 Defensive Burying Test
18.7 Social Interaction
18.8 Separation-induced Ultrasonic ``Distress´´ Vocalization
18.9 Vogel Conflict Test
18.10 Staircase Test
18.11 Hole-Board Test
18.12 Open Field Test
18.13 Conclusion and Future Perspectives
Bibliography
19: Screening Methods forthe Evaluation of AntiparkinsonianDrugs
19.1 Introduction
19.2 In vitro Techniques
19.2.1 Primary Microglial Cultures
19.2.2 Animal Striatal Slices
19.2.3 Dopamine Receptors Assessment by Radioligand Binding
19.2.4 Dopamine Transporter Assessment by Radioligand Binding
19.2.5 Dopamine Release from Synaptosomes
19.2.6 Assessment of Neuroprotective Efficacy
19.3 In vivo Models
19.4 Drug-Induced Animal Models of PD
19.4.1 Reserpine Model of PD
19.4.2 Methamphetamine Model of PD
19.4.3 Neuroleptics Induced PD Models
19.4.4 Cholinomimetics Induced PD Models
19.5 Toxin Induced PD Models
19.5.1 MPTP Models in Monkeys
19.5.2 Rotenone Induced PD Models
19.5.3 Paraquat Model for PD
19.5.4 Maneb-Paraquat Model
19.5.5 6-OHDA (6-Hydroxy Dopamine or Oxdopamine) Model of PD
19.5.6 Manganese
19.6 Genetic Models of PD
19.6.1 α-Synuclein Transgenic Mouse Models
19.6.2 α-Synuclein Transgenic Drosophila Model
19.6.3 Parkin Transgenic Mouse Models
19.6.4 Pink-1 Deleted Mutant Drosophila and Mice Model
19.6.5 DJ-1 Deficient Transgenic Mouse/Drosophila Models
19.6.6 LRRK2 Gene Knock-in Transgenic Mouse
19.6.7 Mitopark Mice
Bibliography
20: Screening Methods for theEvaluation of Drugs for Learning and Memory
20.1 Introduction
20.2 In vitro Methods
20.2.1 In vitro Inhibition of Acetylcholine-Esterase (AChE) Activity in Rat Striatum
20.2.2 In vitro Inhibition of Butyrylcholine-Esterase (BChE) Activity in Human Serum
20.2.3 In vitro Estimation of Presynaptic Release of [3H]- ACh and Other Transmitters from Rat Brain
20.2.4 In-vitro [3H]-Oxotremorine-M Binding to a Muscarinic Cholinergic Receptor in Rat Forebrain
20.2.5 In-vitro [3H]-N-Methylscopolamine Binding in Presence/Absence of Guanylyimidophosphate
20.2.6 Transfected Cell Lines
20.2.7 Evaluation of beta and gamma Secretase Inhibition
20.2.8 Cultures of Rat Hippocampal and Cerebral Cortex Neurons
20.2.9 Study of Field Excitatory Postsynaptic Potential
20.2.10 Other in-vitro Techniques Used for Evaluating Activity in AD
20.3 In-vivo Behavioral Models:Passive (Inhibitory) Avoidance Tests
20.3.1 Step Down Model
20.3.2 Step Through Model
20.3.3 Uphill Avoidance Model
20.3.4 Two-Compartment Test
20.3.5 Trial to Criteria Inhibitory Avoidance
20.4 In-vivo Behavioral Models: Active Avoidance Tests
20.4.1 Runaway Avoidance Test
20.4.2 Shuttle Box (Two-way) Avoidance Test
20.4.3 Jumping (One-way Shuttle) Avoidance Test
20.5 In-vivo Discrimination Learning Tests
20.5.1 Spatial Habituation Learning
20.5.2 Spatial Habituation Learning
20.5.3 Spatial Learning in a Radial Arm
20.5.4 Spatial Learning in the Water Maze (Morris Test)
20.5.5 Visual Discrimination Test
20.5.6 Olfactory Discrimination Test
20.6 In-vivo Experiments in Other Animals
20.6.1 Aversive Discrimination in Chickens
20.6.2 Discrimination Studies in Aged Monkeys
20.7 Genetic Models for Testing Memory and Learning
20.8 Invertebrate Models
Bibliography
21: Screening Methods for the Evaluation of Antianginal Agents
21.1 Introduction
21.2 In-vitro Models
21.2.1 Isolated Heart (Langendorff) Preparation
21.2.2 Gottlieb and Magnus Method (Balloon Method)
21.2.3 Doring and Dehnert (1998) Method
21.2.4 Isolated Rabbit Aorta Preparation
21.2.5 Calcium Antagonism in the Pitched Rat
21.2.6 Relaxation of Bovine Coronary Artery
21.2.7 Coronary Artery Ligation in Isolated Rat Heart
21.2.8 Isolated Heart-Lung Preparation
21.2.9 Plastic Cast Technique in Dogs
21.3 In-vivo Methods
21.3.1 Occlusion of Coronary Artery in Anesthetized Dogs and Pigs
21.3.2 Microspheres Induced Acute Ischemia
21.3.3 Isoproterenol-Induced Myocardial Necrosis in Rats
21.3.4 Stenosis-Induced Coronary Thrombosis Model
21.3.5 Electrical Stimulation-Induced Coronary Thrombosis
21.3.6 Myocardial Ischemic Preconditioning Model
21.3.7 Models of Coronary Flow Measurement
Bibliography
22: Screening Methods for the Evaluation of Antihypertensive Drugs
22.1 Introduction
22.2 Measurement of Blood Pressure in Animals
22.3 Animal Models of Hypertension
22.4 In-Vitro Models of Hypertension
22.4.1 Endothelin Receptor Antagonism in Porcine Isolated Hearts
22.4.2 Monocrotaline Induced Pulmonary Hypertension
22.4.3 ACE inhibition in Isolated Guinea Pig Ileum
22.4.4 Isolated Guinea Pig Atria
22.5 Rat models of Hypertension
22.5.1 Reno-Vascular Hypertension Models in Rats
22.5.1.1 Two Kidney One Clip (2K1C) Hypertension [Goldblatt Hypertension Model]
22.5.1.2 One Kidney One Clip Model (Chronic Renal Hypertension in Rats)
22.5.1.3 Two-Kidney Two Clip Method [Chronic Renal Hypertension in Rats]
22.5.2 Dietary Hypertension Models in Rats
22.5.2.1 Fructose-Induced Hypertension in Rats
22.5.2.2 High Salt-Induced Hypertension in Rats
22.5.3 Neurogenic Hypertension Model in Rats
22.5.4 Endocrine Hypertension
22.5.4.1 DOCA Salt-Induced Hypertension in Rats
22.5.5 Psychogenic Hypertension Models in Rats
22.5.6 Genetic Models of Hypertension in Rats
22.6 Dog Models of Hypertension
22.6.1 Chronic Renal Hypertension
22.6.2 Neurogenic Hypertension
22.7 Monkey Models of Hypertension
22.7.1 Renin Inhibition in Monkeys
22.8 Transgenic Models
22.8.1 Transgenic Rats Overexpressing Mouse Ren-2 Gene [TGR (mRen 2) 27]
22.8.2 Knockout Models
Bibliography
23: Screening Methods for the Evaluation of Antiarrhythmic Drugs
23.1 Introduction
23.2 In Vitro Models
23.2.1 Isolated Guinea Pig Papillary Muscle
23.2.2 Langendorff Technique
23.2.3 Acetylcholine or Potassium Induced Arrhythmias
23.3 In-Vivo Models
23.3.1 Chemically Induced Arrhythmias
23.3.2 Electrically Induced Arrhythmias
23.3.3 Exercise-Induced Ventricular Fibrillation
23.3.4 Mechanically Induced Arrhythmias
23.3.5 Genetically Induced Arrhythmia
Bibliography
24: Screening Methods for the Evaluation of Cardiotonic Drugs
24.1 In vitro and Ex vivoMethods
24.1.1 Ouabain Binding Assays
24.1.2 Isolated Hamster Hearts
24.1.3 Isolated Cat Papillary Muscle
24.2 In Vivo Methods
24.2.1 Mouse Models
24.2.2 Rat models
24.2.2.1 Models of Myocardial Injury
24.2.2.2 Rat Left Coronary ArteryLigation Model
24.2.2.3 Rat Aortic Banding Model (Pressure Overload Model)
24.2.2.4 Dahl Salt-Sensitive Rats
24.2.2.5 Spontaneous Hypertensive Rats and Spontaneous Hypertensive-Heart Failure Rats (SH-HF Rats)
24.3 Large Animal Models
24.3.1 Dog Models
24.3.1.1 Chronic Rapid Pacing Model
24.3.1.2 Volume Overload Model
24.3.1.3 Pressure Overload Model
24.3.1.4 Coronary Artery Ligation and Microembolization Model
24.3.2 Rabbit Models
24.3.2.1 Volume and Pressure Overload Model
24.3.2.2 Tachycardia Pacing Model
24.3.2.3 Doxorubicin Cardiomyopathy Model
24.3.3 Guinea Pig Models
24.3.3.1 Cardiac Insufficiency Model
24.3.3.2 Heart Failure Associated with Sudden Cardiac Death
24.3.3.3 DOCA-Salt Associated Heart Failure
24.3.4 Syrian Hamster Models
24.3.4.1 Cardiomyopathic Syrian Hamsters
24.4 Genetic Mice Models
24.4.1 Myosin Lim Protein Knockout Mice
24.4.2 TNF-Alpha Overexpressing Mice
24.5 Newer Target Identification in CHF and Other Modalities of Treatment
Bibliography
25: Screening Methods for the Evaluation of Antiplatelet Drugs
25.1 Introduction
25.2 Targets for the Antiplatelet Drug
25.3 Antiplatelet Drugs
25.4 Screening for Antiplatelet Drugs
25.4.1 Invitro Screening Method
25.4.1.1 Plate Aggregation in Platelet-Rich Plasma (PRP) or Washed Platelets (WP)
25.4.1.2 Platelet Aggregation by Laser Scattering Mmethod
25.4.1.3 Platelet Aggregation After Gel Filtration
25.4.1.4 Microchannel Array Flow Analyzer (MC-FAN)
25.4.1.5 Platelet Function Aanalyzer-100 (PFA-100)
25.4.2 In-Vivo Models
25.4.2.1 Chemically Induced Thrombosis Model
FeCl3 Induced Thrombosis
25.4.2.2 Photochemical Model for Thrombosis
25.4.2.3 Rose Bengal Induced Thrombosis
25.4.2.4 Laser-Induced Thrombosis Model
25.4.2.5 Arteriovenous Shunt Model for Thrombosis
25.4.2.6 Electrical Induced Thrombosis Model
25.4.2.7 Mechanically Induced Thrombosis Model/Stenosis Induced Thrombosis Model
25.4.3 Zebrafish as a Thrombosis Model
25.4.4 Genetic Models
Bibliography
26: Screening Methods for the Evaluation of Diuretics
26.1 Introduction
26.2 In Vitro Methods
26.2.1 Carbonic Anhydrase Inhibition In Vitro
26.2.1.1 Principle
26.2.1.2 Procedure
26.2.1.3 Evaluation
26.2.2 Patch-Clamp Technique in Kidney Cells
26.2.2.1 Principle
26.2.2.2 Procedure
26.2.2.3 Evaluation
26.2.3 Perfusion of Isolated Kidney Tubules
26.2.3.1 Principle:
26.2.3.2 Procedure
26.2.3.3 Evaluation
26.2.4 Isolated Perfused Kidney
26.2.4.1 Principle
26.2.4.2 Procedure
26.2.4.3 Evaluation
26.3 In Vivo Methods
26.3.1 Diuretic Activity in Rats (LIPSCHITZ Test) (Refer Fig. 26.2)
26.3.1.1 Principle
26.3.1.2 Procedure
26.3.1.3 Evaluation
26.3.2 Saluretic Activity in Rats
26.3.2.1 Principle
26.3.2.2 Procedure
26.3.2.3 Evaluation
26.3.3 Micropuncture Technique in the Rat (Refer Fig. 26.3)
26.3.3.1 Principle:
26.3.3.2 Procedure:
26.3.3.3 Evaluation
26.3.4 Diuretic and Saluretic Activity in Dogs (Refer Fig. 26.4)
26.3.4.1 Principle
26.3.4.2 Procedure:
26.3.4.3 Evaluation:
26.3.5 Clearance Methods (Refer Fig. 26.5)
26.3.5.1 Principle (Refer Table 26.5):
26.3.5.2 Procedure
26.3.5.3 Evaluation:
26.3.6 Stop-Flow Technique (Refer Fig. 26.6)
26.3.6.1 Principle
26.3.6.2 Procedure
26.3.6.3 Evaluation
Bibliography
27: Screening Methods for the Evaluation of Antihyperlipidemic Drugs
27.1 Lacunae with Current Anti-Hyperlipidemics
27.2 How to Choose a Relevant Model for Screening?
27.3 Classification of Screening Methods for Anti-Hyperlipidemias
27.3.1 In-Vitro Screening Methods
27.3.1.1 Inhibition of Isolated HMG-CoA Reductase
27.3.1.2 In Vitro Assay Using Caco-2 Cell Lines
27.3.1.3 ACAT Inhibitory Model
27.3.2 In-Vivo Methods
27.3.2.1 Triton-Induced Hyperlipidemic Rat Model
27.3.2.2 Cholesterol Diet-Induced Hyperlipidemia
27.3.2.3 Chronic Model or High-Fat Diet (HFD) Induced Hyperlipidemia
27.3.2.4 Propylthiouracil (PTU) Induced Hyperlipidemia
27.3.2.5 Fructose Induced Hyperlipidemia in Rats
27.3.2.6 Hypolipidemic Activity in Syrian Hamsters
27.3.2.7 Hereditary Hyperlipidemia in Rats and Rabbits
27.3.2.8 Transgenic Animal Models
Bibliography
28: Screening Methods for the Evaluation of Antiulcer Drugs
28.1 Introduction
28.2 In Vitro Methods
28.2.1 Neutralization Effect on Artificial Gastric Acid
28.2.2 Vatier´s Artificial Stomach Apparatus Model
28.2.3 Fordtran´s Model
28.3 In Vivo Method: Gastric Ulcer Evaluation
28.3.1 Acute Gastric Ulcer Evaluation
28.3.1.1 Rats: Pylorus Ligation Method (Shay Rat Model)
28.3.1.2 Stress Ulcer Model in Rats: Restrain-Induced Ulcers (Immobilization Stress), Hanson and Brodie
28.3.1.3 Stress Ulcer Model in Rats: Cold Water Immersion Restraint (WRS) Stress, Takagi et al
28.3.1.4 Stress Ulcer Model in Rats: Cold Restraint stress, Senay and Levine (1967); Das et al (1993)
28.3.1.5 Stress Ulcer Model in Rats: Stress and NSAID-Induced Ulcers
28.3.1.6 Stress Ulcer Model in Rats: Haemorrhagic Shock-Induced Gastric Ulcers
28.3.1.7 Histamine-Induced Ulcer in Guinea Pigs
28.3.1.8 NSAIDs-Induced Gastric Ulcer
28.3.1.9 Ethanol-Induced Mucosal Damage in Rats
28.3.2 Sub-Acute Gastric Ulcer Evaluation
28.3.3 Chronic Gastric Ulcer: Acetic Acid-Induced Gastric Ulcer
28.4 In Vivo Method: Duodenal Ulcer Evaluation
28.4.1 Cysteamine-Induced Duodenal Ulcers
28.4.2 Chemical-Induced Duodenal Ulcers
Bibliography
29: Screening Methods for the Evaluation of Antidiarrheal Drugs
29.1 Introduction
29.2 In Vitro Methods
29.3 In-Vivo Methods
29.3.1 Chemical-Induced Diarrhoea
29.3.1.1 Castor Oil-Induced Diarrhoea
29.3.1.2 Magnesium Sulphate Induced Diarrhoea
29.3.1.3 Serotonin Induced Diarrhoea
29.3.2 Chemical-Induced Enteropooling
29.3.2.1 Castor Oil-Induced Enteropooling
29.3.2.2 PGE2 Induced Enteropooling
29.3.3 Gastrointestinal Transit Time Using Charcoal Meal
29.3.4 Antidiarrheal Effect in Cecectomized Rats
29.3.5 Antidiarrheal Effect in Cold-Restrained Rats
Bibliography
Untitled
30: Screening Methods for the Evaluation of Antiemetics
30.1 Introduction
30.2 In-Vitro Methods
30.3 In-Vivo Methods
30.3.1 Chemicals/Drugs Induced Emesis Model
30.3.1.1 Apomorphine Induced Emesis
30.3.1.2 Copper Sulphate Induced Emesis
30.3.1.3 Cisplatin-Induced Emesis
30.3.1.4 Cisplatin-Induced Delayed Emesis
30.3.1.5 Methotrexate Induced Delayed Emesis
30.3.1.6 Other models of drug-induced emesis
30.3.2 Emesis Induced by Motion
30.3.3 Emesis Induced by Radiation
Bibliography
31: Screening Methods for the Evaluation of Hepatoprotective Agents
31.1 Introduction
31.2 Classification of Liver Diseases
31.3 Causes of Liver Disease
31.3.1 Hepatotoxins
31.3.2 General Mechanism of Action of Hepatotoxins
31.4 Liver Function Tests
31.5 Hepatoprotective Agents
31.6 Screening Models
31.6.1 In Vitro Models
31.6.2 Ex Vivo Models
31.6.3 In Vivo Models
31.7 Evaluation of Hepatoprotective Effect
31.8 Conclusion
Bibliography
32: Screening Methods for the Evaluation of Antiobesity Drugs
32.1 Introduction
32.2 Animal Models for Inducing Obesity
32.2.1 High Fat Diet-Induced Obesity
32.2.2 Cafeteria Diet-Induced Obesity
32.2.3 Fat or Sugar Choice Diet-Induced Obesity
32.2.4 Meal Feeding-Induced Obesity
32.2.5 Surgically-Induced Hypothalamic Obesity
32.2.6 Ovariectomy-Induced Obesity in Rat
32.2.7 Castration-Induced Obesity in Male Rat
32.2.8 Gold-Thioglucose-Induced Hypothalamic Obesity
32.2.9 Monosodium Glutamate-Induced Obesity
32.2.10 Drug-Induced Obesity
32.3 Genetically Obese Animals
32.4 Uncommon Models Used in Obesity
32.4.1 Obesity Induction in Macaques
32.4.2 Drosophila Model for Obesity
32.5 In-vitro Assay Methods for Evaluation of Anti-Obesity Activity
32.5.1 Measurement of GDP Binding in Rat´s Brown Adipose Tissue
32.5.2 Measurement of Uncoupling Protein-1 in Rat´s Brown Adipose Tissue
32.5.3 β3-Adrenoceptor Binding in the Chinese Hamster Ovary Cells
32.5.4 Determination of mRNA Level Of Leptin in Rat´s Adipose Tissue
32.5.5 Plasma Leptin Level Determination in Rats
32.5.6 Binding Assay of Neuropeptide Y (NPY) Receptor in Pigs
32.5.7 Binding Assay of Orexin-A and Orexin B Receptors In Hamster Ovary Cells
32.5.8 Binding Assay for Galanin Receptors in Rats
32.5.9 Other In-vitro Assays
32.6 In-Vivo Methods for Evaluation of Anti-obesity Activity
32.6.1 Measuring Resting Metabolic Rate in the Mouse
32.6.2 Measurement of Food Consumption in Rats
Bibliography
33: Absorption Studies
33.1 Introduction
33.2 Importance of Drug Absorption
33.3 Classification of Drug Absorption Studies
33.4 In vitro Methods to Evaluate Drug Absorption
33.4.1 Partition Coefficient
33.4.1.1 Log-P
33.4.1.2 Log-D
33.4.2 Parallel Artificial Membrane Permeability Assay (PAMPA)
33.4.3 Brush Border Membrane Vesicles (BBMV)
33.4.4 Isolated Intestinal Cells
33.4.5 Everted Small Intestinal Sac Technique
33.4.6 Everted Sac Modification Technique
33.4.7 Cell Culture Techniques
33.5 In-Vivo Models to Evaluate Drug Absorption
33.5.1 Doluisio Method
33.5.2 Intestinal Loop Technique
33.6 In-Situ Method to Evaluate Drug Absorption
33.7 Conclusion
Bibliography
34: Screening Methods for the Evaluation of Antiasthmatic Agents
34.1 Introduction
34.2 In-Vitro Models
34.2.1 Checking for Smooth Muscle Relaxant Action
34.2.2 Testing Anti-inflammatory Action
34.3 Animal Models
34.3.1 The Murine Allergic Airway Inflammation Model
34.3.2 Other Animal Models
34.4 Measuring Outcomes
34.5 Example of Published Trial Testing Anti-muscarinic in an Acute Model of Asthma
34.6 Example of Published Trial Testing a Chronic Model for Late-Phase Response
Bibliography
35: Screening Methods for the Evaluation of Antitussives and Expectorants
35.1 Introduction
35.2 Cough Induced by Chemical Stimuli
35.2.1 Citric Acid-Induced Coughing in Guinea Pigs
35.3 Cough Induced by Mechanical Stimuli
35.4 Cough Induced by Stimulation of Superior Laryngeal Nerve
35.5 Screening of Expectorants
35.5.1 In Vitro Studies of Mucus Secretion
35.5.2 In Vivo Studies of Mucus Secretion
35.5.2.1 Acute Studies
35.5.2.2 Chronic Cannulation Studies
Bibliography
36: Screening Methods for the Evaluation of Antidiabetic Drugs
36.1 Introduction
36.2 In-Vivo Models for T1DM
36.2.1 Chemical Models for T1DM
36.2.1.1 Streptozotocin-Based model
36.2.1.2 Alloxan-Based Model
36.2.1.3 Other Chemical Models
36.2.2 Hormonal Models for T1DM
36.2.3 Antibody-Mediated Models for T1DM
36.2.4 Microbiological Models for T1DM
36.2.5 Surgical Models for T1DM
36.2.6 Genetic Models for T1DM
36.3 In-Vivo Models for T2DM
36.3.1 Chemical Models for T2DM
36.3.2 Genetic Models for T2DM
36.3.2.1 Monogenic Models
36.3.2.2 Polygenic Models
36.3.2.3 Transgenic and Knockout Models
36.3.2.4 Miscellaneous Models
36.4 Models for Diabetic Complications
36.4.1 Models for Diabetic Nephropathy
36.4.2 Models for Diabetic Retinopathy
36.4.3 Models for Diabetic Neuropathy
36.4.4 Models for Diabetic Cardiac Disease
36.5 In-Vitro Models
Bibliography
37: Screening Methods for the Evaluation of Antithyroid Drugs
37.1 Introduction
37.2 Thyroid Hormones
37.2.1 Historical Assays
37.2.2 In Vitro Tests for Thyroid Hormones
37.2.3 In Vivo Tests for Thyroid Hormones
37.3 Antithyroid Drugs
37.3.1 In Vivo Model for Antithyroid Activity
37.3.2 Antithyroidal Effects in Animal Assays
37.4 Conclusion
Bibliography
38: Screening Methods for theEvaluation of Antifertility Drugs
38.1 Introduction
38.2 Physiology of Female Sexual Organs
38.2.1 Follicular Phase or Proliferative phase
38.2.2 Luteal or Secretory Phase
38.3 Estrous Cycle in Rodents
38.4 Experimental Animal Models
38.4.1 Invivo Methods
38.4.1.1 Methods for Females
Test for Anti-Ovulatory Activity
38.4.2 Test for Estrogenic Activity
38.4.3 Test for Anti-Estrogenic Activity
38.4.4 Test for Progestational Activity
38.4.4.1 Clauberg-McPhail Method
38.4.4.2 Endometrial Carbonic Anhydrase Activity
38.4.4.3 Deciduoma Reaction in Rats
38.4.4.4 Assays causing changes in ovulation
38.4.4.5 Pregnancy Maintenance Test
38.4.5 Test for Anti-Progestational Activity
38.4.6 Test for Anti-Implantation Activity /Abortifacient Activity
38.4.7 Methods for Males.
38.4.8 Test for Androgenic and Anabolic Activity:
38.5 Test for Anti-Androgenic Activity
38.6 Invitro Methods
38.6.1 Methods for Females
38.6.1.1 Test for Estrogenic Activity
38.6.1.2 Techniques used to determine various parameters
38.6.2 Test for Anti-Estrogenic Activity
38.6.3 Test for Progestational Activity
38.6.4 Methods for Males
38.6.4.1 Emergent Spermatozoa Made Non-Functional
38.6.5 Test for Anti-Androgenic Activity
Appendix
Phases of Menstrual Cycle
Vaginal Opening Assay.
Appearance of the vagina in different phases of estrous cycle of a Swiss albino strain mouse. a-Proestrus, b-Estrus, c- Metest...
Bibliography
39: Screening Methods for the Evaluation of the Drugs Acting on Posterior Pituitary, Adrenal Steroid, Testicular, Parathyroid,...
39.1 Screening of Drugs Acting on the Thyroid System
39.1.1 Tadpole Tail Culture Method
39.1.2 Thyroidectomy Model
39.1.3 Iodine Release Inhibition
39.1.4 Anti-Goitrogenic Activity
39.1.5 Reduction in Tensile Strength
39.2 Screening of Drugs Acting on the Androgenic System (Table 39.2)
39.2.1 Sebum Secretion in Rats
39.2.2 Castration Technique
39.2.3 Chicken Comb Model
39.2.4 Nitrogen Retention Assay
39.2.5 Growth of Secondary Sex Organs
39.2.6 Change in Beta-glucuronidase Activity
39.3 Screening of Drugs Acting on the Estrogenic System (Table 39.3)
39.3.1 Castration of Female Rats
39.3.2 Vaginal Cornification
39.3.3 Increased Uterine Weight
39.3.4 Chick Oviduct Method
39.4 Screening of Drugs Acting on Progesterone System (Table 39.4)
39.4.1 McPhail/Clauberg Test
39.4.2 Rat Uterine C3 Model
39.4.3 Rat Decidualization Test
39.4.4 Rat Ovulation Inhibitionspiepr146 model
39.5 Screening of Drugs Acting on Posterior Pituitary/Oxytocin System
39.5.1 Milk Ejection Method
39.5.2 Chicken Blood Pressure Method
39.5.3 Isolated Uterus Method
39.6 Screening of Drugs Acting on the Adrenal System
39.6.1 Screening of Glucocorticoids (Table 39.5)
39.6.1.1 Adrenalectomy in Rats
39.6.1.2 Atrophy of Adrenal, Thymus Gland, and Spleen and Reduced Lymphocyte Count in Rats
39.6.2 Screening of Mineralocorticoids (Table 39.6)
39.6.2.1 Electrolyte Excretion
39.7 Screening of Drugs Acting on the Parathyroid System (Table 39.7)
39.7.1 Serum Calcium Increase Model
39.7.2 Serum Phosphate Decrease Model
39.7.3 cAMP Release Model
Bibliography
40: Screening Methods for the Evaluation of Analgesics, Anti-Inflammatory Drugs, and Antipyretics
40.1 Screening for Analgesic Activity
40.1.1 In Vitro Models for Analgesic Screening
40.1.1.1 3H-Naloxone Binding Assay
40.1.1.2 3H-Dihydromorphine Binding Assay
40.1.1.3 3H-Bremazocine Binding Assay
40.1.1.4 Enkephalinase Inhibition Assay
40.1.1.5 Nociceptin Receptor Binding Assay
40.1.1.6 Cannabinoid Receptor Binding Assay
40.1.1.7 Vanilloid Receptor Binding Assay
40.1.1.8 VIP Receptor Binding Assay
40.1.1.9 Bioassay Techniques for Peripheral Mediators of Central Analgesia
40.1.2 In Vivo Models for Analgesic Screening
40.1.2.1 In Vivo Models for Central Analgesic Activity
40.1.2.2 In Vivo Models for Peripheral Analgesic Activity
40.2 Screening for Anti-Inflammatory Activity
40.2.1 In Vitro Methods for Anti-Inflammatory Screening
40.2.2 In Vivo Methods for Anti-Inflammatory Screening
40.2.2.1 In Vivo Methods for Acute and Subacute Inflammation
40.2.3 In Vivo Methods for Chronic Inflammation
40.2.4 Miscellaneous Models of Anti-Inflammatory Activity
40.3 Screening for Antipyretic Activity
40.3.1 Brewer´s Yeast Suspension Method
40.3.2 Bacterial Lipopolysaccharide Method
40.3.3 Other Less Commonly Used Methods
Bibliography
41: Screening Methods for the Evaluation of Antiglaucoma and Anticataract Drugs
41.1 Introduction
41.2 Glaucoma
41.2.1 Types of Glaucoma
41.2.2 Aqueous Humor Outflow
41.2.3 Intraocular Pressure Measurement
41.2.4 Designing the Study to Test the Drug Efficacy
41.2.5 Animal Models
41.3 Cataract
41.3.1 Animal Models of Congenital Cataracts
41.3.2 Animal Models: Induced Cataract
41.3.3 Hereditary Animal Models
41.3.4 Diabetic-Induced cataracts
41.4 Conclusion
Bibliography
42: Screening Methods for the Evaluation of Drugs Affecting Peripheral Nerve Functions
42.1 Screening of Skeletal Muscle Relaxants (Neuromuscular Blocking Agents)
42.1.1 Scope and Limitations of Neuromuscular Screening Tests
42.1.2 In-Vitro Screening Tests
42.1.2.1 Isolated Nerve-Muscle Preparations
42.1.2.2 Single Muscle Fiber Study
42.1.2.3 Ionophoretic Testing Method
42.1.2.4 Human In Vitro Preparations
42.1.3 Screening Tests in Anesthetized Animals
42.1.4 Screening Tests in Intact Non-Anesthetized Animals
42.1.4.1 Inclined Plane Method
42.1.4.2 Rotarod Method
42.1.4.3 Testing of Righting Reflexes
42.1.4.4 Head Drop Technique
42.1.4.5 Others
42.1.5 Screening Tests in Anesthetized Human Beings
42.1.6 Screening Tests in Intact Non-Anesthetized Human Beings
42.2 Screening of Local Anesthetics
42.2.1 Screening for Surface Anesthesia
42.2.2 Screening for Infiltration Anesthesia
42.2.3 Screening for Conduction Anesthesia
42.2.4 Screening for Spinal and Epidural Anesthesia
Bibliography
43: Mutagenic Toxicity Testing
43.1 Introduction
43.2 History of Genotoxicity Testing
43.3 Introduction to Genetic Toxicology Assays
43.3.1 Metabolic Activation System
43.3.2 Genotoxicity Testing Endpoints
43.4 Predictors of Genotoxicity: Structural Alerts and In Silico Assays
43.5 Assays for DNA Damage and Repair
43.5.1 Comet Assay for DNA Strand Breakage
43.5.2 DNA Repair Assays
43.5.2.1 In Bacteria
43.5.2.2 Unscheduled DNA Synthesis in Cultured Mammalian Cells and Intact Animals
43.5.3 Assays Based on DNA Stress Response Mechanisms
43.5.3.1 GreenScreen Assay or GADD45a-GFP Assay
43.6 Gene Mutation Assays in Prokaryotes
43.7 Assays in Nonmammalian Eukaryotes
43.7.1 Sex-Linked Recessive Lethal Test
43.7.2 Plant Assays
43.7.3 Mitotic Recombination in Yeast
43.8 Mammaliangene Mutation Assays
43.8.1 In Vitro Assays for Gene Mutation in Mammalian Cells
43.8.2 In Vivo Assays for Gene Mutation in Mammalian Cells
43.8.2.1 Mouse Spot Test
43.8.2.2 In Vivo HPRT Assay
43.8.2.3 Pig-a Mutation Assay
43.8.3 Transgenic Assays
43.9 Mammalian Cytogenetic Assays
43.9.1 Chromosome Aberrations
43.9.1.1 In Vitro Mammalian Chromosome Aberration Test
43.9.2 In Vivo Mammalian Chromosome Aberration Test
43.9.3 Micronucleus Assay
43.9.4 Sister Chromatid Exchange
43.9.5 Aneuploidy
43.10 Germ Cell Mutagenesis Assays
43.10.1 Mouse Specific-Locus Test
43.10.2 Cytogenetic Assays
43.10.3 Mouse Heritable Translocation Assay
43.10.4 Dominant Lethal Assay
43.10.5 Assays for DNA Damage
43.11 Regulatory Guidelines for Genotoxicity Testing
43.12 Conclusion
Bibliography
44: Screening Methods for the Evaluation of Anticancer Drugs
44.1 Introduction
44.2 In-Vitro Methods
44.2.1 Functional Assays (Cell Viability Assays)
44.2.2 Non-Functional Assays (Morphological Assays)
44.2.2.1 Assessment of Mode of Cell Death
44.2.2.2 Assessment of Cell Death Markers
44.2.2.3 Assessment of Genotoxicity of Compounds
44.2.3 Molecular Techniques in In-Vitro Screening
44.3 In-Vivo Screening Methods for Anticancer Drugs
44.3.1 Chemical Carcinogen Models
Box 44.1. Procarcinogen DMH Is Most Commonly Used for Inducing Colorectal Tumors in Rats and Mice
44.3.2 Viral Infection Models
44.3.3 Methods Involving Cell Line/Tumor Pieces Implantation
44.3.4 Genetically Engineered Mouse Models
Bibliography
45: Screening Methods for the Evaluation of Drugs for Benign Prostatic Hyperplasia
45.1 Introduction
45.2 Pathophysiology and Pharmacology of BPH in Brief
45.3 In Vitro Models
45.4 In Vivo Models
45.4.1 Testosterone Induced Benign Prostatic Hyperplasia in Rats
45.4.2 Testosterone + Estrogen Induced Benign Prostatic Hyperplasia in Rats
45.4.3 Sulpiride-Induced BPH in Rats
45.4.4 Bladder Outlet Obstruction of Hemorrhagic Cystitis by Cyclophosphamide in Rats
45.4.5 Single Dose of Cyclophosphamide Induced Hemorrhagic Cystitis in Rats of Bladder Outlet Obstruction Model
45.4.6 Partially Ligated Urethra of Bladder Outlet Model
45.4.7 Intraprostatic Injection Model
45.5 Advantages of In Vivo Models
45.6 Limitations of In Vivo Models
45.7 Ex Vivo Models
45.7.1 Isolated Prostate Gland Contractility Model (Flow Chart Given Below)
45.7.2 Isolated Detrusor Muscle and Urethra Contractility Model
45.8 Advantages of Ex Vivo Models
45.9 Limitations With the Use of Ex Vivo Studies
45.10 Transgenic Models
45.10.1 Probasin Prolactin Transgenic Mouse Model of Benign Prostate Hyperplasia
45.10.2 MMTV (Murine Mammary Tumor Virus) -Int2 (Proto-Oncogene) Transgenic Mice Model
45.10.3 MMTV-Keratinocyte Growth Factor (fgf7)-Induced Hyperplasia of the Transgenic Model
45.11 Advantages of Transgenic Models
45.12 Limitations of Transgenic Models
45.13 Xenograft Models
45.14 Advantages With the Use of Xenograft Models
45.15 Limitations with the Use of Xenograft Models
45.16 Spontaneous BPH Model
45.17 Conclusion
Bibliography
46: Screening Methods for the Evaluation of Drugs Acting on Autonomic Nervous System
46.1 Introduction
46.2 Screening of Unknown Substance for ANS Activity (Irwin Method)
46.3 Sympathetic System
46.3.1 In-Vitro Assays
46.3.2 Isolated Organ Models
46.3.3 In Vivo Models
46.4 Parasympathetic System
46.4.1 In Vitro Model
46.4.2 Isolated Organ Model
46.4.3 In Vivo Model
Bibliography
47: Screening Methods for the Evaluation of Anti-Infective Agents
47.1 Screening of Antifungal Drugs
47.2 Screening of Anthelminthic Drugs
47.3 Screening of Antibacterial Drugs
47.4 Screening of Antiviral Drugs
47.4.1 HIV-Related Screening Methods
47.4.2 Non-HIV-Related Screening Methods
Bibliography
48: Preclinical Toxicity Studies
48.1 Introduction
48.2 Acute Toxicity Study
48.2.1 Aim of Acute Toxicity Study
48.2.2 Definition
48.2.3 Animal Selection
48.2.4 Number of Animals
48.2.5 Sex of Animals
48.2.6 Age of the Animals
48.2.7 General Care of the Animals
48.2.8 Administration of the Testing Compound
48.2.9 Route of Drug Administration
48.2.10 Selection of Dose Levels
48.2.11 Observation Period
48.2.12 Calculation of Medial Lethal Dose 50 (LD50)
48.2.12.1 Staircase or Up-Down Method of LD50 Calculation
48.2.12.2 Karber´s Method for Calculation of LD50
48.2.12.3 Miller and Tainter´s Graphical Method of Calculation of LD50
48.2.12.4 LD50 Calculation by Lorke´s Method
48.2.13 Summary of Acute Toxicity Studies
48.3 Sub-Acute Toxicity Studies (Repeated 14-28Days Toxicity Study)
48.3.1 Aim
48.3.2 General Considerations
48.3.3 Calculation of Sub-Acute Lethal Dose
48.4 Sub-Chronic (90Days Repeated Dose) Toxicity Study
48.4.1 Aim
48.4.2 General Considerations
48.4.3 Calculation of Sub-Chronic Lethal Dose
48.5 Chronic (180Days - 1Year Repeated Dose) Toxicity Study
48.5.1 Aim
48.5.2 General Considerations
48.5.3 Calculation of Chronic Lethal Dose
48.5.4 Calculation of NOAEL and LOAEL
48.5.5 Calculation of Reference Concentration (RfC) or Reference Dose (Rfd) or Acceptable Daily Intake (ADI) from NOAEL or LOA...
48.6 Reproductive Toxicity Studies
48.6.1 General Considerations
48.6.2 Protocols
48.6.3 Assessment of Male Reproductive Toxicity
48.6.4 Assessment of Female Reproductive Toxicity
48.6.5 Endpoints and Indices in Reproductive Toxicity Studies
48.7 Local Toxicity Studies
48.7.1 Dermal Toxicity Studies
48.7.2 Ocular Toxicity Studies
48.7.3 Inhalational Toxicity Studies
Bibliography
49: Ethical Issues in Animal Research
49.1 Introduction
49.2 Terms Related to Research in Animals: The Definitions
49.3 Animal Experimentations: To Do or Not to Do?
49.4 The R Principles: 3Rs, 4Rs, and so on
49.4.1 Replacement: The 1st R
49.4.2 Reduction Alternative: The 2nd R
49.4.3 Refinement Alternative: The 3rd R
49.4.4 Miscellaneous Principles
49.5 Alternatives to Animal Experimentations
49.6 Evolution of Ethics in Animal Research
49.7 Organisation for Economic Co-operation and Development (OECD) Guidelines
49.8 Compendium of CPCSEA
Box 49.1: Ethical principles adopted by CPCSEA for use of animals in scientific experiments
49.9 Guide for the Care and Use of Laboratory Animals, 8th edition
49.10 Animal Experimentation in India and Elsewhere: What Is the Current Status?
49.11 Conclusion
Bibliography
50: Good Laboratory Practice (GLP)
50.1 Introduction
50.2 Principles of Good Laboratory Practice
50.2.1 Test Facility: Organization and Personnel
50.2.1.1 Test Facility Management
50.2.1.2 Study Director
50.2.1.3 Principal Investigator
50.2.1.4 Study Personnel
50.2.2 Quality Assurance Programme (QAP)
50.2.3 Facilities
50.2.4 Apparatus, Material, and Reagents
50.2.5 Test Systems
50.2.6 Test and Reference Items
50.2.7 Standard Operating Procedures (SOPs)
50.2.8 Performance of the Study
50.2.9 Reporting of Study Results
50.2.10 Storage and Retention of Records and Materials
50.3 GLP Advantages and Drawbacks
50.3.1 Advantages of GLP
50.3.2 Drawbacks of GLP
50.4 National Good Laboratory Practice (GLP) Compliance Monitoring Authority (NGCMA)
Bibliography
Part II: Biochemical Pharmacology
51: Basic Principles and Applications of Simple Analytical Methods
51.1 Introduction
51.2 Qualitative Drug Tests
51.3 Analytical Techniques
51.3.1 Titrimetric Techniques
51.3.2 Spectrophotometry and Beer Lambert´s Law
51.4 Sample Preparation for Drug Analyses
Bibliography
52: Principles of Quantitative Estimation of Drugs, Endogenous Compounds, and Poisons-1
52.1 Chromatography
52.2 Types of Chromatography
52.3 Paper Chromatography
52.3.1 How It Works
52.4 Thin-Layer Chromatography (TLC)
52.4.1 How It Works
52.5 Gas Chromatography (GC)
52.5.1 Working Principle
52.5.2 Mobile Phase
52.5.3 Stationary Phase
52.5.4 Samples for GC Analysis
52.5.5 Instrumentation
52.5.6 Gas Chromatography: Mass Spectrometry (GC-MS)
52.6 Liquid Chromatography (LC)
52.6.1 Modern Liquid Chromatography
52.6.2 Working Principle
52.6.3 Instrumentation
52.6.3.1 Solvent Delivery System
52.6.3.2 Sample Injectors
52.6.3.3 Columns
52.6.3.4 Detectors
52.6.4 Chromatogram Output
52.6.5 LC-MS
52.7 Mass Spectrometry
52.8 Working Principle and Instrumentation
52.8.1 Sample Introduction
52.8.2 Ionization
52.8.3 Mass Analyzers
52.8.4 Tandem MS
52.8.5 Detector
52.9 Enzyme-Linked Immunosorbent Assays
52.9.1 Working Principle
52.9.2 Types of ELISA
52.9.2.1 Direct ELISA
52.9.2.2 Indirect ELISA
52.9.2.3 Sandwich ELISA
52.9.2.4 Competitive/Inhibition ELISA
52.9.3 Enzymes Commonly Used in ELISA
52.9.4 Different Steps Performed in ELISA
Bibliography
53: Principles of Quantitative Estimation of Drugs, Endogenous Compounds, and Poisons-2
53.1 Colorimetry
53.1.1 Introduction
53.1.2 Principle
53.1.3 Instrumentation
53.1.4 Methodology
53.1.5 Applications
53.1.5.1 Applications of Colorimetry in Pharmacology
53.1.5.2 Advantages of Colorimetry
53.1.6 Limitations of Colorimetry
53.2 Spectrophotometry
53.2.1 Introduction
53.2.2 Principles of Spectrophotometry
53.2.3 Instrumentation
53.2.4 Types of Spectrophotometers
53.2.5 Sample Measurement
53.2.6 Applications of Spectrophotometer
53.2.7 Drawbacks or Disadvantages of Spectrophotometer
53.2.8 Safety Considerations
53.3 Photometry
53.3.1 Introduction
53.3.1.1 Key Terms in Photometry
53.3.2 Principle
53.3.3 Instrumentation
53.3.4 Methodology
53.3.5 Applications of Photometry
53.3.6 Advantages
53.3.7 Limitations
53.4 Flame Photometry
53.4.1 Introduction
53.4.2 Principle
53.4.3 Instrumentation
53.4.4 Working of Flame Photometry
53.4.5 Methodology
53.4.6 Applications of Flame Photometry
53.4.7 Advantages
53.4.8 Disadvantages
53.5 Fluorimetry
53.5.1 Introduction
53.5.2 Principles of Fluorimetry
53.5.3 Instrumentation
53.5.4 Fluorescence Measurement
53.5.5 Applications of fluorometry
53.6 Radioimmunoassay (RIA)
53.6.1 Background
53.6.2 Introduction
53.6.3 Principle
53.6.4 Types of RIA
53.6.5 Materials Required
53.6.6 Methodology
53.6.7 Applications
53.6.8 Advantages
53.6.9 Limitations
53.7 Nuclear Magnetic Resonance (NMR) Spectroscopy
53.7.1 Introduction
53.7.2 NMR Principle
53.7.3 NMR: Sample handling and working
53.7.4 Instrumentation
53.7.5 NMR Interpretation
53.7.6 Applications of NMR Spectroscopy
Bibliography
Untitled
54: Plant Extraction Methods
54.1 Introduction
54.2 Classification
54.3 Traditional Extraction Methods
54.3.1 Maceration
54.3.2 Soxhlet Extraction
54.3.3 Reflux Extraction
54.3.4 Decoction
54.3.5 Infusion
54.3.6 Percolation and Re-percolation
54.4 Novel/Modern (Non-conventional) Plant Extraction Methods
54.4.1 Microwave Extraction
54.4.2 Ultrasound-Assisted Extraction (Sonication Extraction)
54.4.3 Supercritical Fluid Extraction
54.4.4 Pressurized Liquid/Solvent Extraction
54.4.5 Pulsed Electric Field Extraction
54.4.6 Vibrocavitation Homogenizer Extraction
54.4.7 Enzyme Assisted Extraction
54.4.8 Hydro/Steam Distillation
54.4.9 Two Phase Extraction
54.5 Conclusion
Bibliography
Part III: Research Methodology
55: Literature Search
55.1 Introduction
Box 55.1. Various Reasons for Conducting a Literature Search
55.2 Types of Medical Literature
55.3 Types of Literature Search
55.3.1 Electronic Research Databases
55.4 Methodology of Literature Search
55.4.1 Translating Research Question to Keywords (Refer Fig. 55.5)
55.4.2 Searching all the Sources: Keyword Search
55.4.3 Techniques to Improve Literature Search
55.4.4 Keeping a Record of Search Activity
55.4.5 Reviewing and Refining the Search Results by Further Literature Search
Bibliography
56: Study Designs
56.1 Introduction
56.2 Evidence-Based medicine
56.3 Concept of Variables
56.4 Types of Research
56.5 Importance of Study Designs
56.6 Types of Study Designs
56.6.1 Descriptive Studies
56.6.2 Analytical Studies
56.6.3 Interventional Studies
56.7 Miscellaneous Study Designs
56.8 Levels of Evidence
Bibliography
57: Ethical Issues Related to Medical Research on Human Participants
57.1 Introduction
57.1.1 Definitions
57.1.2 Respect for Autonomy
Box 57.1. Essential Components of Informed Consent Document
57.1.3 Nonmaleficence
57.1.4 Beneficence
57.1.5 Justice
57.2 History of Medical Research Ethics: How It Started?
57.2.1 Declaration of Helsinki
57.2.2 Council for International Organizations of Medical Sciences (CIOMS) Guidelines
57.2.3 The Common Rule (The Final Rule)
57.2.4 Indian Context
57.3 Ethics Review Committees
57.4 Ethics in Different Contexts
57.4.1 Underprivileged Populations
57.4.2 Publication Ethics
57.5 Medical Research Ethics: What is the Current Status?
Box 57.2. Placebos in Clinical Research
57.6 Conclusion
Bibliography
Part IV: Biostatistics
58: Basic Principles and Application of Statistics in Drug Research
58.1 Why to Use Statistics?
58.2 Uses of Statistics in Drug Research
58.3 Some Important Terms
Box 58.1
58.4 Important Principles for the Application of Statistics in Clinical Research
Bibliography
59: Calculation of Basic Statistical Parameters
59.1 Introduction
59.2 Variable
59.2.1 Types of Variables
59.2.1.1 Quantitative Variable (Numeric Variable)
Discrete Variable
Continuous Variable
59.2.1.2 Qualitative Variable (Categorical Variable)
Nominal Variable
Ordinal Variable
59.3 Measures of Central Tendency
59.3.1 Mean
59.3.2 Median
59.3.3 Mode
59.4 Measures of Dispersion
59.4.1 Standard Deviation
59.4.1.1 Interpretation of Standard Deviation
59.4.2 Variance
59.4.3 Range
59.4.4 Interquartile Range
59.4.4.1 Interpretation of IQR
59.4.5 Normal Distribution Curve
59.4.5.1 Few Characteristics of the Normal Distribution Curve
59.4.6 Summary Measures for Qualitative or Categorical Data
59.4.6.1 Ratio
59.4.6.2 Rate
59.4.6.3 Proportion
59.5 Inferential Statistics
59.5.1 95% Confidence Interval
Bibliography
60: Hypothesis Testing
60.1 Introduction
60.2 Inferential Statistics and Its Domains
60.3 Formulating a Research Question and a Hypothesis
60.4 Null and Alternative Hypothesis
60.5 Type I and Type II Errors
60.6 Steps in Hypothesis Testing
60.6.1 Example of Hypothesis Testing
60.7 Interpreting the p-Value
60.7.1 Problems in Interpretation of p-Value
60.8 Confidence Interval
60.9 Multiple Hypothesis Testing
60.10 Bayesian Hypothesis Testing
60.11 Conclusion
Bibliography
61: Parametric Tests
61.1 Introduction
61.2 The Basic Assumptions Underlying Parametric Tests are as Follows (Box. 61.1)
Box 61.1. Basic Assumptions Underlying Parametric Tests
61.3 General Steps in Performing any of the Parametric Tests
61.4 Selection of Appropriate Parametric Test
61.5 Parametric Testing Methods
61.5.1 T-Test
61.5.1.1 One Sample t-Test
61.5.1.2 Independent Sample t-Test/unpaired t-Test/Student´s t Test
61.5.1.3 The Paired-Samples t-Test
61.5.2 Analysis of Variance (ANOVA)
61.5.2.1 One Way ANOVA Test
61.5.2.2 Two Way ANOVA Test
61.5.2.3 Repeated Measures ANOVA
61.5.3 Pearson´s Correlation
61.5.4 Regression
61.6 Advantages of Parametric Test Over Non-parametric Test
61.7 Circumstances Where Parametric Tests Cannot Be Used
Bibliography
62: Non-parametric Tests
62.1 Introduction to Non-parametric Statistics
62.2 Scope of Non-parametric Tests
62.3 Steps in Performing Non-parametric Tests
62.4 Non-parametric Testing Methods
62.4.1 Sign Test
62.4.2 Wilcoxon Signed-Rank Test
62.4.3 Friedman Test
62.4.4 Goodman Kruskal´s Gamma Test
62.4.5 Kruskal-Wallis Test
62.4.6 Mann-Kendall Trend Test
62.4.7 Mann Whitney U-Test
62.4.8 Mood´s Median Test
62.4.9 Macnemar´s Chi-Squared Test
62.5 Advantages of Non-parametric Tests
62.6 Disadvantages of Non-parametric Tests
Bibliography
63: Common Errors in Using Statistical Tools and Data Presentation
63.1 Introduction
63.2 Errors Related to Sample Size
63.2.1 Manipulation of Confidence Interval (1-α) and Power (1-beta) Values to Achieve Low Sample Size
63.2.2 Extrapolating Conclusions for an Objective From the Sample Size Estimated for Different Objectives
63.2.3 Missing Drop-Out Cases in Sample Size Calculation
63.3 Errors Related to Data Handling
63.3.1 Unnecessary Conversion of Continuous Data to Categorical Data
63.3.2 Setting a Groundless Cut-off Value During Categorization
63.3.3 Unnecessary Data Transformation
63.4 Errors Related to Statistical Tests
63.4.1 Choosing Correct Statistical Tests
63.4.2 Violating the Assumptions of Statistical Test
63.5 Errors in Data Representation
63.5.1 Errors in Data Tabulation
63.5.2 Errors in Graphs
63.5.3 Errors in Descriptive Data Summarization
63.5.3.1 Errors in P-Value Expression
Bibliography