Ijpi-07 Sesha Madhavi

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International Journal of Pharmaceutical Invention

September 2012, Volume 2(8) IJPI 20

EFFECT OF VERAPAMIL HYDROCHLORIDE ON

ACETYLCHOLINE INDUCED CONTRACTION IN FROG RECTUS

ABDOMINIS MUSCLE

K. KARTHIGADEVI*, O. MADHAVI USHARANI, I. SESHA MADHAVI,

CH.V.R.TANUJA, S. KAVIMANI

Department of Pharmacology, College of Pharmacy,

Mother Theresa Post Graduate and Research Institute of Health Sciences (Govt. of

Puducherry Institution), Indria Nagar, Gorimedu, Puducherry-605006, India.

E-mail: [email protected]

Keywords: Verapamil Hydrochloride, Calcium channel, Frog Rectus Abdominis Muscle,

Right Hand Side Shift.

INTRODUCTION

Verapamil Hydrochloride is an oral and

parenteral calcium-channel blocking

(CCB) agent belonging to the

phenylalkylamine class. Verapamil

Hydrochloride is useful for the treatment

of angina, hypertension, and for

supraventricular tachyarrhythmias. It is

considered a class IV antiarrhythmic agent

and Verapamil Hydrochloride is more

effective than digoxin for controlloing

Verapamil Hydrochloride is an oral and parenteral calcium-channel blocking (CCB)

agent belonging to the phenylalkylamine class. Verapamil Hydrochloride is useful for

the treatment of angina, hypertension, and for supraventricular tachyarrhythmias. It is

considered a class IV antiarrhythmic agent and Verapamil Hydrochloride is more

effective than digoxin for controlling ventricular rate in patients with atrial

fibrillation. Skeletal muscle contraction is explained by the excitation-contraction

coupling and is dependent on the calcium induced release of calcium from the

sarcoplasmic reticulum by Ca2+ release channel due to the previous influx of

Ca2+

through Dihydropyridine receptor present on the T-tubule which is due to the

conduction of action potential on T-tubule. So, an attempt is made to find out the

effect of Verapamil Hydrochloride on acetylcholine induced contraction in Frog

Rectus Abdominis Muscle. Maximum response produced by acetylcholine in the

absence of the blockers was taken as 100% and against that the % response was

calculated. Verapamil Hydrochloride produced significant dose dependant inhibition

of acetylcholine induced contraction in CRC.


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K. KARTHIGADEVI et al ISSN:22771220


than digoxin for controlling ventricular

rate in patients with atrial fibrillation.

Verapamil Hydrochloride was synthesized

in 1962 and in 1981, became the first

calcium channel blocker to be approved

by the FDA. Verapamil Hydrochloride

inhibits the influx of extracellular calcium

across the myocardial and vascular

smooth muscle cell membranes. It exerts

its activity at the membrane surface of

arterial smooth muscle cells and withinconductile and contractile tissue in the

myocardium, but the serum calcium levels

remain unchanged.[ 1 ]

The calcium channel antagonists are a

mature group of drugs directed at

cardiovascular diseases including

hypertension, angina, peripheral vascular

disorders and some arrhythmic conditions.

Their sites and mechanism of actions have

been well explored over the past two

decades and their interactions at the

1subunit of L-type channels (Cav1.1-1.4)

have made them valuable molecular tools

for channel classification and localization.

With the realization that other members of

the voltage-gated calcium channel family

exist Cav 2.1-2.3 and Cav 3.1-3.3

considerable effort has been directed to

drug discovery at these channel types

where therapeutic prospects exist for a

variety of disorders including pain,

epilepsy, affective disorders,

neurodegenerative disorders, etc. In

contrast to the situation with the L-type

channel antagonists success in developing

small molecule antagonists of therapeutic

utility for these other channel types has

thus far been lacking. The reasons for this

are explored and potential new directions

are indicated including male fertility, bone

growth, immune disorders, cancer and

schistosomiasis.[ 2 ]

To explore and to investigate for new

calcium channel blockers first it wasthought to have an extensive knowledge

over calcium regulation by different

calcium channels and their structures to

have a look on binding sites of drugs.

The concentration of cytosolic free

calcium ([Ca2+

]i)is critically important for

the control of many essential cellular

responses. Changes in [Ca2+

] can control

specialized functions like excitability,

contraction, or exocytosis, due to which

neurotransmitter release takes place, while

also regulating universal cellular activities

such as metabolism and gene expression.

In most cells, elevations([Ca2+

]i)in arise

from [Ca2+

] entry via Ca2+

channels in the

surface membrane, or Ca2+

discharge from

internal stores, or both. Mechanisms that

tend to return ([Ca2+

]i) toward its low

resting value include extrusion of [Ca2+

]

across the surface membrane or

resequestration by internal [Ca2+

] stores

and by Ca2+ pump or Na+-Ca2+

exchangers.[ 3 ]


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MATERIALS AND METHOD

A pithed frog was laid on its back on the

frog dissecting board. The skin on the

abdomen was removed and the rectus

abdominis muscle was exposed. The one

side of the rectus muscle was dissected

from the pelvic girdle to the pectoral

girdle. The muscle was then pinned to the

cork so as to keep it stretched to its

normal length and soaked in frog ringer

solution. A thread was sewn through eachend and the muscle was mounted in organ

bath containing frog ringer solution at

room temperature. The concentration

response curve was obtained as described

by Kulkarni (2009). The experiment was

repeated in each set of preparation in

presence of Verapamil Hydrochloride (0.4

and 0.8 g/ml).The tissue was irrigated

for 20 minutes. Maximal response

produced by acetyl choline in the absence

of the blocker was taken as 100% and

against that the percentage response was

calculated.[ 4 ]

RESULTS

Verapamil Hydrochloride at various test

doses (0.4 and 0.8g/ml) produced

significant dose dependent inhibition of

acetylcholine induced contractions inconcentration response curves. 640g/ml

of acetylcholine was required to produce

100% of response where as in the

presence of Verapamil Hydrochloride

(0.4 and 0.8g/ml), the same

concentration of acetylcholine produced

92% and 66% ofresponses respectively.

Fig 1: Effect of Verapamil Hydrochloride on Ach Induced Contractions in Frog Rectus

Abdominis Muscle.


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Fig 2: The graph represents the log dose- response curves of Acetylcholine (100 g/ml)

in presence of various concentrations of Verapamil Hydrochloride (0.4 and 0.8g/ml).

The progressive shift to the right hand side was noted indicating the antagonistic effect

of Verapamil Hydrochloride.

DISCUSSION

Any muscle contraction is possible only

through the cellular calcium signaling and

cellular calcium regulation is the

substantial preoccupation of the cell.

Heilbrunn described the calcium is the

most ubiquitous intracellular signaling

molecule, which serves both physiological

and pathologically. It plays an important

role in signal transduction pathways,

where it acts as a second messenger, in

neurotransmitter release from neurons by

exocytosis, contraction of all muscle cell

types. Many enzymes require calcium

ions as a cofactor; those of the blood-

clotting cascade being notable examples.

Extracellular calcium is also important for

maintaining the potential difference across

excitable cell membranes, as well as

proper bone formation.[ 3,5 ]

Regulation of Ca2+

is under very tight and

dynamic control in all major cell

compartments (cytoplasm, nucleus,

endoplasmic reticulum, mitochondria). In

each compartment, this control is

achieved through the interplay of

transmembrane entry, extrusion systems

(channels, exchanger, transporters) and of

buffering systems (Ca2+

binding protein).

It is this role of calcium as an

information-bearing second messenger

together with the control processes that

make possible the existence of calcium

controlling drugs. A triumvirate of

hormones-vitamin D, parathyroid

hormone and calcitonin- that serve to


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regulate calcium absorption, calcium

excretion and bone deposition and

resorption (remodelling), achieves this

control.[6 ]

THE KEY ROLE OF Ca2+

and Ca2+

CHANNEL IN CELLULAR

SIGNALING

Ca2+

channels allow passage of Ca2+

ions

into the cytoplasm through a selective

pore which is opened in response to

depolarization of the cell membrane. TheCa

2+flux creates a net inward,

depolarizing current and the resulting

accumulation of Ca2+

in the cytoplasm can

act as a chemical trigger for secretion of

hormones and neurotransmitters,

contraction of muscle and a variety of

other Ca2+

sensitive events. Thus, upon

sensing membrane potential changes,

Ca2+

channels simultaneously generate an

electrical signal while directly creating an

intracellular chemical messenger. This

dual ability is unique among the family of

ion channels and allows the Ca2+

channel

to play a variety of roles in excitation-

secretion and excitation-contraction

coupling.

Excitation-Contraction Coupling

Skeletal muscle contraction is explained

by the excitation-contraction coupling and

is dependent on the calcium induced

release of calcium from the Sarcoplasmic

reticulum by Ca2+ release channel (RyR)

due to the previous influx of Ca2+

through

Dihydropyridine Receptor (DHPR)

present on the T-tubule which is due to

the conduction of action potential on T-

tubule. The inhibitory effect of Verapamil

Hydrochloride and diltiazem may be due

to the blockade of receptor operated

calcium channel and voltage dependent

calcium channel.Sarcoplasmic reticulum

and single channel studies have provided

evidence that phenylalkylamine calcium

antagonists inhibit calcium releasethrough the sarcoplasmic reticulum

calcium channel/ryanodine receptor. This

action has not been observed with

dihydropyridine calcium antagonists.[ 7 ]

CONCLUSION

The acetylcholine induced contractions in

isolated skeletal muscle preparation was

effectively antagonised by Verapamil

Hydrochloride. It also showed dose

dependent antagonistic effect. The log

dose response curves of acetylcholine in

presence of Verapamil Hydrochloride was

shifted to the right hand side indicating

the antagonistic effect. The antagonistic

effect of Verapamil Hydrochloride

(phenylalkylamine) may be due to the

blocking of both L-type Ca2+

channel

(DHPR) and Ryanodine Ca2+

sensitive

channel (RyR) which are involved in the

skeletal muscle contractions.

REFERENCES1. http://www.druginfosys.com


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2. TriggleDJ, Calcium ChannelAntagonists: Clinical uses past,

present and future, Biochem

Pharmacology, 74(1), 2007 June 30,

1-9.

3. Richard W. Tsien, Rogery Y. Tsien,Calcium Channels, Stores and

Osallation, Annual Review, Cell

Biology, 6, 1990: 715-60.

4. S.K.Kulkarini, Handbook ofExperimental Pharmacology, 3

rd

edition, 2009, VallabhPrakashan

Publications

5. http://en.wikipedia.org/wiki/Calcium_channel_blockers

6. Stefan I McDonough, CalciumChannel Pharmacology, Springer,

2004.

7. Ricardo Zucchi, Effect of gallopamilon excitation-contraction coupling,

General Pharmacology: The vascular

system, 27(5), July 1996, 749-7

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