1. Dr. Mohd Nazam Ansari

2.  Isolated tissue preparations are commonly used to study the effects of drugs on specific type of receptors. These preparations are also used:  For bioassay of drugs,  Characterization of specific receptor or its subtypes,  To determine concentration response curve of an agonist,  To study antagonism of drug and in new drug discovery.

4.  The in vitro isolated preparation represents an isolated organ or a piece of living tissue from a freshly killed animal.  Optimum ionic environment  Adequate supply of nutrition and oxygen  And a stable temperature  These basic requirements should be provided if one has to maintain the isolated tissue in living state.

5.  Physiological Salt Solutions  Instrumentation  Procedures and drugs – to render the animal unconscious  Tissue: isolated or whole

6.  Optimal pH 7.0 – 7.2  Commonly used PSS are:  Frog Ringer (for heart, rectus abdominis and other preparations of frog)

7.  Tyrode (for guinea pig ileum, rat ileum, rabbit ileum etc.)  De Jaon (for rat uterus)  Kreb’s solution (for rat fundus strip, tracheal preparations, vas deferens) etc.

8. IngredientsConcentration on gms/l Frog ringerTyrodeKreb’s bicarbonateDe jalon NaCl KCl CaCl 2 NaH 2 PO 4 KH 2 PO 4 MgSO 4 MgCl 2 Glucose NaHCO 3 6.5 0.14 0.12 0.05 - 1.5 0.4 8.0 0.2 0.18 0.1 - 0.1 1.0 5.9 0.35 0.28 - 0.16 0.11 - 2.0 2.1 9.0 0.35 0.003 - 0.5 NaCl: To maintain isoosmolarity, isotonicity, excitability and contractility KCl: To maintain ionic balance CaCl 2 : To maintain contractility NaH 2 PO 4 & KH 2 PO 4 : Act as a buffer MgSO 4 & MgCl 2 : To stabilize the preparation and hence reduce the spontaneous activity Glucose : To provide energy NaHCO 3 :To provide alkaline pH

11.  The apparatus providing the basic requirements of life to the tissue and facilitating the record of response.  Outer water bath of perspex or glass  Inner organ bath (single or multiple)-glass (15-100 ml): Organ tube  Tissue holder cum Aeration (oxygen) tube  Glass coil connected to organ bath, Marriott's bottle: Reservoir for PSS  Electric heater  Preheating coil

12.  Oxygen or carbogen (5% CO 2 + 95% Oxygen)  The purpose of aeration are:  Supply of oxygen  Uniform distribution of drug due to bubbling  Maintenance of pH specially in Kreb’s solution where amount of NaHCO 3 used is more

13.  It is an instrument on which physiological responses are recorded.  Heavy base  Base hoofs (legs) with adjustable leveling screws  Gear rod  Vertical shaft  Drum Cylinder  Brass or iron It has 8 variable speeds. The speed is measured in mm per second. Eight variable speeds are 0. 12, 0. 25, 1. 25, 2. 5, 12. 5, 25, 320 and 640 mm per second.

15.  Isometric contractions: change in tension (force). E.g. spring lever  Isotonic contractions: change in length at uniform tension. E.g. isotonic frontal writing lever (light aluminum or stainless steel rod)

16.  Glazed paper  A mixture of benzene and kerosene (1:9)

17.  A solution of resin (colophony) in methylated spirit. This solution is prepared by dissolving 150 gm of resin in 2 Lit of spirit.  Recording of responses on drum cylinder without smoking (Sketch-Pen Tip)

18.  For slowly contracting tissue e.g. rectus abdominis muscle it is 60-90 sec.  For fast contracting tissue e.g. Guinea pig ileum it is 30 sec.

19.  Step by step  Setup the assembly first  Assembly should be on right hand side and drum should be on left hand side  The drum should move from right to left or away from the writing point  The speed of drum should be minimum  The writing lever should be horizontal and the writing point should touch the drum.  All tracing should begin after the overlap of the paper.  Always start with the minimum concentration of drug.  Write the name & amount of drug. Name of experiment and student with student ID and signature of teacher before varnishing.