NEUROTOXINSPRATHAMESH KUDALKAR

13FET1011

What are Neurotoxins?

Neurotoxins are the substances that are poisonous or destructive to the nerve cells.

The term is also used for classifying endogenous compounds which when present in higher concentrations may prove neurologically toxic.

They affect the functioning of developing as well as mature nervous tissues.

Neurotoxins can be of plant, animal, mineral or atmospheric origins.

Neurotoxins inhibit neuron control over ion concentrations across cell membrane or communication between neurons across a synapse.

COMMON NEUROTOXINS

Botulinum Toxin Lead

Teradotoxin Ethanol

Tetanus Toxin Glutamate

Nitric Oxide Tetraethylammonia

Chlorotoxin Arsenic

Conotoxin Mercury

Botulinum Toxin

Botulinum Toxin (BTX) is a group of neurotoxins consisting

of eight distinct compounds, referred to as BTX-

A,B,C,D,E,F,G,H

Botulinum Toxin (BoNT) causes a rare disease termed as

Botulism.

It is the most potent of all the known poisons. 1g is

sufficient to kill approximately 30 billion mice.

LD50 value for humans is 0.2ng/kg.

It is produced by bacterium Clostridium Botulinum.

Botulinum Toxin

Biology of C.Botulinum

Gram Positive

Rod Shaped

Anaerobic

Spore forming

Motile

Large

Growth Conditions

pH (>4.6)

Water activity (>0.93)

Anaerobic

Moderately High Salt

(5.5%)

Temperature(35-37

degree Celsius)

Botulinum Toxin Toxin

Seven serotypes (A, B, C, D, E, F and G) of botulism

are recognized, based on the antigenic specificity of

the neurotoxin produced by each strain.

Types A, B, E and F cause human botulism

Types C and D cause most cases of botulism in

animals. As they are highly potent toxins only a very

small amount (1ng/kg body weight) is necessary for

severe symptoms and even death.

The toxins are heat labile and can be destroyed by

high and uniform heat at 950C for 15 minutes or

boiling for 5 minutes.

Botulinum Toxin

Groups I II III IV

Neurotoxin Types A, B, F B, E, F C, D G

Gene Location Chromosome Chromosome Bacteriophage

Activity Proteolytic Non Proteolytic Non Proteolytic Proteolytic

Optimum Temp. 35-40 18-25 15 ND

Minimum aw 0.94 0.97 ND ND

Minimum pH 4.6 5.0 ND ND

D100 (min) 25 <0.1 0.1-0.9 0.8-1.12

D121 (min) 0.1-0.2 <0.001 ND ND

STRUCTURE The toxin is in the form of a non-covalently bound

complex that contains several nontoxic proteins that

consist of hemegglutinin and nonhemegglutinin, and

weighs 150 kDa

In order for this polypeptide molecule to become

toxic, it is cleaved by a protease at one-third the

distance from the N terminus.

The exact enzyme that performs this function has still

yet to be been determined.

This action yields two fragments: a smaller, lighter

fragment weighing 50 kDa and a heavier fragment

with a larger weight of 100 kDa

Proteolytic activity is located at the N-terminal end of

the light chain of botulinum toxin

MECHANISM OF ACTION Outside the body.

Food

Heat

MECHANISM OF ACTION

Inside the body

The neurotoxin targets the body’s peripheral nervous system.

After entering the body, it is transported by blood stream to the nerve cells

They pass through the presynaptic membrane of motor nerve terminals

and enter into the neuron cell by endocytosis.

The heavier chain forms a channel within the membrane in which the lighter chain can then pass through into the cytoplasm.

The botulinum toxin produces specified cleaving proteases that allow

the pathogen to successfully attach to the synaptic vesicles.

MECHANISM OF ACTION The carboxy-terminal domain of the heavy chain recognizes a specific binding

site, while the nitrogen-terminus transports the lighter chain into the nerve cytosol

The lighter chain contains metalloproteases that target SNARE proteins involved in

controlling the exocytosis machinery.

Neurotoxin types depend upon the characteristic of lighter chain.

SNARE proteins are group of proteins which facilitate the release of acetylcholine

from Synaptic vesicles.

Type B,D,F,G cleaves Synaptobrevin

Type A,C,E cleaves SNAP-25

Type C cleaves Syntaxin

Thus membranes of vesicles carrying acetylcholine and presynaptic membranes

do not fuse, in turn blocking the exocytosis of neurotransmitter acetylcholine

MECHANISM OF ACTION Acetylcholine plays an essential role in the body

It is responsible for regulating the somatic nervous system,

which controls the voluntary movements of the skeletal

muscles, and is the only of its kind.

If acetylcholine is absent, muscle contraction cannot take

place resulting in regional flaccid paralysis of muscle.

It affects the cardiovascular as well as respiratory muscles

Death is caused mainly due to respiratory failure.

Exposure and transmission Foodborne botulism

Infant botulism

It occurs when infants ingest Clostridium botulinum spores, which germinate into

bacteria that colonize in the gut and release toxins.

In most adults and children older than about six months, this would not happen

because natural defense that develops over time prevent germination and

growth of the bacterium.

Wound botulism

Wound botulism is rare and occurs when the spores get into an open wound and

are able to reproduce in an anaerobic environment.

Other types of intoxication

Water borne

Airborne

FOOD SOURCES The spores are heat-resistant (killed at 1150C) and can

survive in foods that are incorrectly or minimally

processed.

Spores do not germinate in the presence of nitrate (250

ppm).

Foods most commonly associated are low acid

vegetables (green beans, corn, spinach, asparagus,

pepper and mushrooms) and fruits (figs and peaches)

and also fermented, improperly cooked and smoked

fish, meat, poultry and fish eggs in hermatically sealed

containers.

SYMPTOMS & TREATMENT At the initial stage (generally 12h to 36h, but can be 2h), some

gastrointestinal symptoms (nausea, vomiting, diarrhea and

constipation) are evident.

Neurological symptoms appear in a short time particularly when the

amount of toxin consumed is more, which includes blurred or

double vision, difficulty in swallowing, breathing and speaking,

dryness of the mouth, and paralysis of different involuntary muscles

that spreads to the lung and heart.

The person who has been intoxicated by botulinum toxin should be

given supportive care for breathing like Ventilator

Equine antitoxin must be administered within 18 hrs

Even if extremely small dosage is ingested, antitoxins must be taken

immediately or it may lead to paralysis.

DETECTION OF TOXIN Mouse bioassay was the earliest method used for detection of

Botulinum toxin

Modern Methods:

ELISA

PCR

HPLC

DNA Microarrays

Real time PCR

GC

TETRODOTOXIN Tetrodotoxin, frequently

abbreviated as TTX, is a extremely

toxic neurotoxin.

It is found mainly in fishes such

as pufferfish, porcupinefish, ocean

sunfish etc.

TTX is found in

the liver, gonads, ovaries,

intestines, and skin of these fish.

LD50 value for humans is 25mg/kg

Tetrodotoxin intoxication is found

mainly in Japan

POISONING The toxin can enter the body by ingestion, injection, or

inhalation, or through abraded skin.

The mechanism of toxicity is through the blockage of fast

voltage-gated sodium channels.

These are required for the normal transmission of signals

between the body and brain.

As a result, TTX causes paralysis of voluntary muscles (including

the diaphragm and intercostal muscles, stopping breathing),

loss of vagal regulation of heart rate (causing it to increase to

around 100bpm), and loss of sensation etc.

SYMPTOMS & TREATMENT The diagnosis of pufferfish poisoning is based on the observed

symptomology and recent dietary history.

Symptoms develop within 30 minutes after ingestion and include

paresthesia of the lips and tongue is followed by hyper-salivation,

sweating, headache, weakness, lethargy, incoordination, tremor, paralysis etc

Death usually occurs in 4-6 hrs. if proper treatment is not provided.

Therapy is supportive.

If ingested, treatment can consist of emptying the stomach, feeding

the victim activated charcoal to bind the toxin, and taking standard

life-support measures to keep the victim alive until the effect of the poison has worn off.

CONOTOXINConotoxins are found mainly in creatures having shells

like snail, oyster and sometimes even crabs.

It inhibits the activity of Ca channel. Some Conotoxins

were found to have different activity.

Cases of Conotoxin intoxication have not been reported

yet.

Nowadays, Conotoxins are permitted by FDA for their

use as Pain Killers.

Not much research has been done on Conotoxins.

TOXIC METALS Metal contamination in food is easily possible during cultivation,

harvesting and processing

Mercury ( Dimethyl mercury) is the most potent toxin ever.

Aluminium and Mercury are capable of inducing CNS damage by

migrating into brain by crossing brain, bone barrier

Arsenic is widely found in the soil. It is destructive towards

cytoskeleton

Lead is a potent neurotoxin whose toxicity has been recognized for

at least thousands of years.

Lead results in increased absorption of calcium in cell leading to

apoptosis (cell death)

GLUTAMATE & NITRIC OXIDE Glutamate and Nitric Oxide are widely used in food as an additives.

Glutamate and Nitric Oxide are part of CNS and hence termed as

endogenous neurotoxins.

Nitric oxide (NO) is commonly used by the nervous system in inter-

neuron communication and signaling.

Glutamate is its functions as an excitatory neurotransmitter.

It is only when these endogenous compounds become highly

concentrated that they lead to dangerous effects.

Increased concentrations lead to DNA damage, swelling and

apoptosis.

Also, NO is carcinogenic.

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