NON-EXPERIMENTAL PRE AND POST-LABORATORY DISCUSSION

Diuretics

are drugs that increase the rate of urine flow (diuresis) and the excretion of sodium (natriuresis) and water.

Diuretics

These are primarily used to treat conditions such as

hypertension, heart failure, and edema

Diuretics

MECHANISM OF ACTION

• Inhibits the reabsorption of Na+ and Cl- at different segments of the renal tubule, leading to increased solute concentration in the filtrate and subsequent increased water excretion.

LOOP DIURETICS (Furosemide)

• Highly efficacious

• Inhibit the Na+, K+, and Cl- in the thick ascending limb of the Loop of Henle

Loop of Henle:

site with high Na+ reabsorption capacity

THIAZIDE DIURETIC  (Hydrochlorothiazide)

Inhibit the Na+ and Cl- cotransporter in the distal convoluted tubule

POTASSIUM - SPARING DIURETICS (Spironolactone, Amiloride)

Act on the collecting duct.

Spironolactone

Aldosterone antagonist

Amiloride

• blocks epithelial Na+ channels.

(diuretics) BASIC PHARMACOLOGIC ASSAY

The main objective is to quantify and compare the diuretic effect of different agents.

(diuretics) BASIC PHARMACOLOGIC ASSAY

• test animal

• Rats or mice

• standardized by weight and fasting status.

(diuretics) BASIC PHARMACOLOGIC ASSAY

• pharmacologic assay

• Saline-loaded model (Lipschitz method)

• Control: saline only

• Test Group: saline + drug

Saline-loaded model (Lipschitz method):

• Animals are fasted, hydrated with a standard volume of saline (to ensure adequate urine output), and divided into control and test groups

(diuretics) BASIC PHARMACOLOGIC ASSAY

• observation & measurement

• Animals are placed in metabolic cages (individual housing with collection devices). 

• The volume of urine excreted over a specific time (e.g., 5 hours) is collected and measured. 

• Electrolyte concentration (Na+, K+, Cl−) in the urine is measured

(diuretics) BASIC PHARMACOLOGIC ASSAY

• calculation

• Diuretic Index (ratio of urine volume of test group to control group)

• Natriuretic Index (ratio of Na+ excretion in test group to control group)

are calculated and compared

Diuretic Index

(ratio of urine volume of test group to control group)

Natriuretic Index

(ratio of Na+ excretion in test group to control group)

(diuretics) BASIC PHARMACOLOGIC ASSAY

• equipment

• Metabolic cages

• oral gavage needles/feeding tubes 

  • for drug administration

• measuring cylinders 

  • for urine volume 

• Flame Photometer or Ion Selective Electrodes 

  • for electrolyte analysis such as Na+ and K

LOCAL ANESTHESIA

Drugs that produce a reversible loss of sensation (analgesia) in a localized area of the body without causing loss of consciousness. They are classified into esters and amides 

LOCAL ANESTHESIA

• esters

procaine

LOCAL ANESTHESIA

• amides

lidocaine

LOCAL ANESTHESIA

• moa

Work by reversibly blocking voltage-gated sodium channels in the neuronal membrane.

LOCAL ANESTHESIA: basic pharmacologic assay

• test animal

• Typically amphibians (e.g., Frog, Toad) for peripheral nerve block studies

• Guinea Pigs/Rabbits for corneal/skin surface anesthesia.

LOCAL ANESTHESIA: basic pharmacologic assay

• pharmacologic assay

• Frog Sciatic Nerve Block (Plexus Anesthesia): 

LOCAL ANESTHESIA: basic pharmacologic assay

• observation & measurement

• Time of Onset (time from application to complete loss of reflex) and

• Duration of Action (time from complete block to full recovery of reflex) are recorded

LOCAL ANESTHESIA: basic pharmacologic assay

• other assays

• Infiltration Anesthesia (e.g., Guinea Pig Skin Wheal):

• Corneal Anesthesia (e.g., Rabbit Eye): 

LOCAL ANESTHESIA: basic pharmacologic assay

• equipment

• Dissecting instruments

• Stopwatch

• stimulator 

  • for reflex testing

• Sterile syringes/needles

• Appropriate restraints/animal holders

Infiltration Anesthesia (e.g., Guinea Pig Skin Wheal)

• Measures loss of sensation in a localized skin area. 

• Frog Sciatic Nerve Block (Plexus Anesthesia): 

• The sciatic nerve is exposed, and the drug solution is applied directly.

  • The time to loss of a reflex (e.g., withdrawal from a painful stimulus) and the time to recovery are measured to assess potency and duration

Corneal Anesthesia (e.g., Rabbit Eye)

• Measures the time taken to abolish the corneal reflex (blinking response to light touch)

ANTIPYRETICS

Agents that reduce an elevated body temperature (fever), such as Paracetamol (Acetaminophen) and Non-Steroidal Anti-Inflammatory Drugs (NSAIDs) like Aspirin and Ibuprofen.

Fever is caused by pyrogens (like bacterial toxins), which trigger the release of

pro-inflammatory cytokines (IL-1, IL-6, TNF-alpha). These cytokines stimulate the production of Prostaglandin E2 (PGE2) in the anterior hypothalamus, which acts as a pyrogen to raise the thermoregulatory set point.

Antipyretics primarily work by

inhibiting the enzyme Cyclooxygenase (COX), which is responsible for the synthesis of prostaglandin E2 (PGE2) from arachidonic acid.

NSAIDS

• Inhibit:

  • both COX-1 and COX-2

  • COX-2 selectively

PARACETAMOL

has a less clear central mechanism but is thought to primarily inhibit COX in the CNS

ANTIPYRETICS: basic pharmacologic assay

• The objective is to induce fever in an animal model and assess the drug's ability to reduce the elevated temperature.

ANTIPYRETICS: basic pharmacologic assay

• test animal

• Rats or Rabbits

• standardized by weight

ANTIPYRETICS: basic pharmacologic assay

• pharmacologic assay

Pyrogen-Induced Fever Model:

ANTIPYRETICS: basic pharmacologic assay

• procedure

1. Animals are divided into control (pyrogen + vehicle) and test groups (pyrogen + drug).

2. Basal rectal temperature is recorded before pyrogen and drug administration.

3. Temperature is monitored at regular intervals (e.g., every 30-60 minutes) for several hours after drug administration

ANTIPYRETICS: basic pharmacologic assay

• observation & measurement

Maximum temperature reduction and duration of antipyretic effect are determined. The drop in rectal temperature is compared between control and test groups

ANTIPYRETICS: basic pharmacologic assay

• equipment

• Rectal Thermometer

  • accurate, digital thermometer designed for small animals 

• syringes/needles 

  • for drug and pyrogen administration)

• animal restraints/holders.

Pyrogen-Induced Fever Model: 

• Fever is induced by administering a pyrogen, such as a suspension of Brewer's yeast (subcutaneously) or Lipopolysaccharide (intravenously). 

• This leads to an increase in body temperature 12-18 hours post-administration.