Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Preclinical Assessment of Potential Central Adverse Effects Department of Pharmacology and Toxicology Faculty of Pharmacy Cairo University Biostatistics and Biological Standardization of Drugs (306) Safety pharmacology Cardiovascular system Central nervous system Respiratory system Secondary organ systems Safety During the drug discovery phase, it is obligatory to assess potential adverse effects of any drug candidate on the cardiovascular system, the central nervous system, the respiratory system and some secondary organ systems in experimental animals before the start of clinical trials (humans) Centrally acting drugs …are capable of passing the blood brain barrier The blood brain barrier is formed of capillary endothelial cells connected by tight junctions that do not exist in normal circulation. That is why the blood brain barrier is highly selective permeable barrier that allows the diffusion of only water, some gases, lipid soluble molecules, glucose and amino acids that are crucial to neural function. Centrally acting drugs cross the blood brain barrier to exert … Therapeutic benefits • • • • • • Antipsychotics Antidepressants Anticonvulsants Sedatives Hypnotics Analgesics Adverse effects • • • • • • • • • Amnesia Convulsion Depression Cognitive impairment Insomnia Involuntary movement Motor incoordination Nausea … Central adverse effects are due to … changes in the structure and/or function of neurons or glial cells in the nervous system Interfering with synthesis, storage, metabolism, release, degradation, uptake, or receptors of metabolism synthesis storage release receptor action reuptake • • • • • • • • • • Glutamate Acetylcholine γ-aminobutyric acid Glycine Dopamine Serotonin Norepinephrine Histamine Opioid peptide and/or Endocannabinoids CNS safety pharmacology Category A/core battery studies are obligatory to be performed at the early stage of drug discovery. They are typically simple tests that can be performed rapidly in a routine fashion. They include: • Functional observational battery (FOB) • Spontaneous motor activity • Locomotor coordination • Convulsive threshold • Interaction with hypnotics • Pain threshold and • Body temperature Category B/supplementary studies include cognitive function, electrophysiology examination, drug dependence and other tests. Category B tests are carried out when necessary. Category A/core battery studies In this practical session, we will screen for potential adverse effects of some drugs on the CNS using modified FOB, spontaneous motor activity and locomotor coordination tests. Drugs are injected 15 min prior to performing the tests and compared with a vehicle control group 1- Functional observational battery (FOB) FOB is used to characterize different neurological functions. It includes observational assessments (innate behaviors, no need to be taught, require little interaction between the observer and the animal) and manipulative tests (require an interaction between the observer and the animal). Observational assessments include Manipulative tests include • Spontaneous activity levels • Neuromuscular tests • Reactivity • Sensory responses • Gait • Posture • Involuntary motor movements • Clinical signs Observational assessments I- Spontaneous activity levels Observe the animal’s spontaneous activity in its home cage. Limit observation to 5 sec or less Rank the level of spontaneous activity displayed in the cage 1 = No activity (animal may be asleep or sitting motionless) 2 = Slight (animal may move its head or body, just a very few times) 3 = Moderate (animal moves about some) 4 = Active (animal moves more actively around cage) 5 = High activity (animal moves about rapidly) Observational assessments II- Reactivity: Observe the level of excitability or resistance of the animal in response to handling and/or removal from the home cage Rank the level of reactivity/excitability using scoring criteria such as: 1= Low (no resistance, easy to hold or pick up) 2= Moderately low (slight resistance) 3= Moderately high (some squirming or moving around) 4= High (excited, squirming, twisting) 5= Very high (aggressive actions e.g. biting, tail and throat rattling) Observational assessments III- Gait: Look at the movement of all four limbs in relation to one another and to the saggital plane of the body Rank the degree of gait abnormality, using a scale such as: 1 = No abnormality 2 = Slight abnormality 3 = Moderate abnormality 4 = Severe abnormality Describe the gait, for example: Ataxia (uncoordinated, staggering, wobbly gait) Hind limbs show exaggerated, overcompensated, or splayed movements Feet (primarily hind feet) point outward from body Forelimbs drag and/or show abnormal positioning Walking on toes Observational assessments IV- Posture: Refers to the placement of the body and spinal curvature Rank the degree of posture abnormality, using a scale such as: 1 = No abnormality 2 = Slight abnormality 3 = Moderate abnormality 4 = Severe abnormality Describe postural alterations using terms such as: Completely flattened, pelvis flat on surface Pelvis low, dragging somewhat Hunched, back raised up Observational assessments V- Involuntary motor movements : Record the occurrence or absence of • Tremors (repetitive contractions/relaxations of major muscle groups) • Fasciculations/twitches (localized muscle contraction) • Convulsions • Sterotypy (repetition of specific gestures o movements) or • Bizarre behaviors (behaviors not normally seen in the animal e.g. straub tail (tail is stiff and is held in a vertical position). Observational assessments VI- Clinical signs : Examine the animal for • Lacrimation • Salivation • Hair coat characteristics (color/staining, alopecia, and piloerection) • Ocular abnormalities (corneal opacity or cloudiness , eyeball appears to bulge) and • Muscle tone or mass (hypotonia or hypertonia) Manipulative tests VII- Neuromuscular tests Indicate the presence or absence of the righting reaction. Hold the animal in a supine position (lying with the face up) on a surface. Quickly release the animal. The animal should immediately flip to resume a normal standing posture with the head turning over first, followed by the forelimbs, and then hind limbs Manipulative tests VIII- Sensory responses Visual approach response: Approach the animal at nose level with the end of a blunt object, such as a pen or pencil. Hold the stimulus ∼3 cm from the face for ∼4 sec to give the animal time to make a response. Rank the magnitude of response to the stimulus: 1 = No reaction or response 2 = Slight or sluggish reaction (flinch or startle as evidence of perception) 3 = Obvious reaction (locomotor orientation as evidence of perception) 4= Clear reaction or response (more intense startle or locomotion) 5= Exaggerated reaction (may jump, bite, or attack Manipulative tests VIII- Sensory responses Somatosensory response: Coming in from the side, touch the rump of the rat gently with a blunt object, such as a pen, pencil, or brush. Use a brief touch (1 to 2 sec) that is deliberate but not too forceful. Rank the magnitude of response to the stimulus: 1 = No reaction or response 2 = Slight or sluggish reaction (flinch or startle as evidence of perception) 3 = Obvious reaction (locomotor orientation as evidence of perception) 4= Clear reaction or response (more intense startle or locomotion) 5= Exaggerated reaction (may jump, bite, or attack Manipulative tests VIII- Sensory responses Auditory response: Position a sound stimulus e.g. metal clicker, finger snap, or hand clap ∼5 cm above the back of the animal (i.e. outside the field of view). Make a sudden sound. Rank the magnitude of response to the stimulus: 1 = No reaction or response 2 = Slight or sluggish reaction (flinch or startle as evidence of perception) 3 = Obvious reaction (locomotor orientation as evidence of perception) 4= Clear reaction or response (more intense startle or locomotion) 5= Exaggerated reaction (may jump, bite, or attack Manipulative tests VIII- Sensory responses Test tail-pinch response: Pinch the tail ∼1 cm from the tip. The very tip of the tail is relatively insensitive. Pinch hard enough to produce a reliable response in control animals, and be consistent in the intensity of the pinch from test to test. Rank the magnitude of response to the stimulus: 1 = No reaction or response 2 = Slight or sluggish reaction (flinch or startle as evidence of perception) 3 = Obvious reaction (locomotor orientation as evidence of perception) 4= Clear reaction or response (more intense startle or locomotion) 5= Exaggerated reaction (may jump, bite, or attack 2- Spontaneous motor activity Can be quantified by manual counting of crossed squares in the open field apparatus The open-field apparatus consists of a square arena of adequate size surrounded by walls to prevent the animal from escaping. The floor is divided into equally spaced squares. The open field apparatus should be located in a quiet room with controlled temperature and ventilation and lowlevel lighting to reduce anxiety. The apparatus should be cleaned prior to the test of each animal to reduce olfactory confounding factors 2- Spontaneous motor activity A. Administer the test drug or the vehicle at an appropriate time prior to placing the mouse into the center of the open field. B. Record the number of squares crossed by the mouse, the number of rearings (when the front legs of the animal are lifted completely off the surface, or when the animal places its front legs on the side of the open-field apparatus) or any stereotype or bizarre behavior within 10 min. Test drug Number of squares crossed Number of rearings Stereotype or bizarre behavior Vehicle 3-Neuromuscular coordination The ability of a rodent to maintain balance and keep pace with a rotating rod has been used to assess motor function. The rotarod consists of a circular rod about 75 cm long, divided into 6 sections to allow simultaneous testing of 6 mice; the rod is attached to a motor rotating the rod at a constant or increasing speed. The rod is placed about 50 cm above a platform to discourage the animals from jumping off the rod. 3-Neuromuscular coordination A. Animals are prescreened; only those that are able to remain on the revolving rod for at least 3 min are used for screening test substances B. Administer the test drug or the vehicle at an appropriate time prior to placing the mouse on the rotarod and then start the motor and timer. C. Record the latency to fall from the rotarod for each mouse. It is generally preferred to take the mean of at least two to three measures from each animal. D. Calculate the percent of animals falling from the rotarod within 3 min Test drug Latency to fall Percent of animals falling within 3 min Vehicle