ENDOCRINE PHYSIOLOGY

Organization of endocrine system

• The endocrine system consists of various endocrine glands
and neurosecretory cells located in the hypothalamus. The
neurosecretory cells of hypothalamus secrete certain
neurohormones called releasing and inhibitory factors which
influence the secretion of hormones from other endocrine
glands.
• Certain other substances act as neurotransmitters in the
brain influence the secretion of neurosecretory cells of
hypothalamus. The environmental factors through these
neurotransmitters influence the whole endocrine system.
The various endocrine glands present in the body are:
1. Pituitary gland (hypophysis).
• Pituitary gland is also known as hypophysis, which in Greek
means undergrowth of the brain. It has two main parts:
adenohypophysis and neurohypophysis.
• Adenohypophysis secretes growth hormone (GH) or
somatotropins, follicle-stimulating hormone (FSH),
luteinizing hormone (LH), prolactin, thyrotropin or thyroid-
stimulating hormone (TSH) and corticotropin or
adrenocorticotropic hormone (ACTH).
• The neurohypophysis stores the antidiuretic hormone (ADH)
or vasopressin and oxytocin synthesized by the
hypothalamus.
2. Thyroid gland.
• The thyroid gland is present in the neck in front of
trachea. It has two lobes and an isthmus (bridge)
connecting the lobes. It secretes thyroxine (T4) and
triiodothyronine (T3).
• The C cells or parafollicular cells which are scattered in
the spaces between the follicles of the thyroid gland
secrete calcitonin.
3. Parathyroid glands.
• These are four in number; very small glands situated
behind the lobes of the thyroid gland and secrete
parathormone.
4. Adrenal glands.
These are situated on the upper poles of the two
kidneys, hence also called suprarenal glands. The outer
cortex region of the adrenal glands secretes cortisol,
aldosterone and sex steroids, and the inner medullary
region secretes catecholamines (adrenaline and
noradrenaline).
5. Pancreatic islets (islets of Langerhans).
These are small groups of cellswhich secrete insulin,
glucagon and somatostatin.
6. Gonads.
• These include ovaries in females and testes in males.
• The ovaries secrete oestrogens and progesterone
(female sex steroids), and testes secrete male sex
hormone (testosterone).
7. Pineal gland.
• It is a small gland present in the roof of third ventricle
in the brain. It secretes melatonin and other biogenic
amines.
8. Placenta.
• During pregnancy, the placenta secretes various
hormones like human chorionic gonadotropin (HCG),
oestrogen, progesterone, somatotropins and relaxin.
9. Gastrointestinal mucosa also secretes various
hormones collectively known as gastrointestinal (GIT)
hormones, e.g. gastrin, secretin, cholecystokinin-
pancreozymin (CCK-PZ), etc.
10. Kidneys.
• In addition to their renal functions, the kidneys secrete
erythropoietin, prostaglandins and 1,25-
dihydroxycholecalciferol, and also help in activation of
angiotensinogen.
11. Atrial muscle cells
• These secrete atrial natriuretic peptides (ANP) and many
other peptides.
12. Skin.
• This is also considered to act as an endocrine structure by
producing vitamin D, which is now considered to be a
hormone.
Hormones: Definition and classification
• The word hormone is derived from the Greek word
hormaein, which means to execute or to arouse.
• In the classic definition, hormones are secretory
products of the ductless glands which are released in
catalytic amounts into bloodstream and transported
to specific target cells (or organs), where they elicit
physiologic, morphologic and biochemical responses.
• In reality, the requirement that hormones be secreted
into the bloodstream is too restrictive, because they
can also act locally.
• Therefore, the chemical messengers that perform hormonal functions
are defined as
1. Endocrine hormones
• These include the chemical messengers whose function is the
transmission of a molecular signal from a classic endocrinal cell
through the bloodstream to a distant target cell .
2. Neurocrine hormones.
• Nervous communication involves the release of chemical messengers
from nerve terminals, which may reach their target cells via one of
three routes:
• Neurotransmitters can be released directly into the intercellular
space, cross the synaptic junction and inhibit or activate the
postsynaptic cell, e.g. acetylcholine and norepinephrine.
• Neural signals can be transferred via gap junction, which is a
membrane stabilization between the nerve cells, between
nerve terminals and endocrine cells and between endocrine
cells.
• Neurohormones or peptides are released from a
neurosecretory neuron into the bloodstream and then
carried to a distant target cells . Example of such neurocrine
substances are oxytocin and antidiuretic hormone (ADH).
The effector sites of neurohormones are not always
endocrine cells.
3. Paracrine hormones. These chemical messengers,
which after getting secreted by a cell, are carried over
short distance by diffusion through the interstitial
spaces (extracellular fluid) to act on the neighbouring
different cell types as or a regulatory substance e.g in
islets of Langerhans, somatostatin secreted by the delta
cells acts on the alpha and beta cells.
4. Autocrine hormones. These refer to those chemical
messengers which regulate the activity of neighbouring
similar type of cells . Examples of autocrine hormones
are prostaglandins.
Different types of hormones (by their mechanism of action) are: A, endocrine
hormone; B, neurocrine hormone; C, paracrine hormone; and D, autocrine hormone .
• Note. It is important to note that according to the route by which it is
transmitted, the same chemical messenger may act as endocrinal
hormone (bloodstream conveyance), or as a paracrine, or as an
autocrine hormone (local conveyance), e.g. insulin secreted by beta
cells in the pancreatic islets may act as:
• Endocrine hormone. When released into bloodstream, insulin acts on
the adipose tissue, muscle, liver and brain to regulate energy stores,
carbohydrate, fat and protein metabolism.
• Paracrine hormone. Insulin, when released into the islets interstitial
fluid, inhibits the neighbouring alpha cells of the islets.
• Autocrine hormone. The insulin released in the islets interstitial fluid
can regulate growth and function of beta cells themselves as they
possess insulin receptors.
Classification of hormones
A. Depending upon the chemical nature
1. Amines or amino acid derivatives, e.g.
• Catecholamines (epinephrine and norepinephrine),
• Thyroxine (T4) and triiodothyronine (T3)
2. Proteins and polypeptides
• i. Short-chain polypeptides include: - Posterior pituitary
hormones - Antidiuretic hormone (ADH) and - Oxytocin.
• ii. Long chain polypeptides include: - Insulin, - Glucagon, -
Parathormone and - Other anterior pituitary hormones.
3. Steroid hormones:
• These include
• Glucocorticoids,
• Mineralocorticoids,
• Sex steroids and
• Vitamin D.
B. Depending upon the mechanism of action
• Group I hormones
• These act by binding to intracellular receptor and mediate
their actions via formation of a hormone receptor complex.
These include steroid, retinoid and thyroid hormones.
• Group II hormones
• These involve second messenger to mediate their effect.
Depending upon the chemical nature of the second
messengers, group II hormones are further divided into four
subgroups A, B, C and D
Types of Group II Hormones Based on the Chemical Nature of
Second Messenger Involved in their Mechanism of Action
 1. Amines or amino acid derivatives Synthesis.
• These hormones include catecholamines, thyroid hormone and, which
are derived from the amino acid tyrosine.
• Thyroid hormones are derived from two iodinated tyrosine residues.
Thyroid hormones are the only substances in the body that contain
iodine. These are synthesized in the cell cytoplasm.
• Storage and release
• Catecholamines are stored in secretory granules inside the cytoplasm of
chromaffin cells.
• Secretion occurs when the membrane of granule fuses with the plasma
membrane, causing the granular contents to be extruded into the
circulation.
• Thyroid hormones are stored outside the follicular cells in the form of
thyroglobulin. Following endocytosis and proteolysis of thyroglobulin,
thyroid hormones are secreted into the bloodstream by simple diffusion.
2. Protein and polypeptide hormones Synthesis
• The peptide and protein hormones are composed of chains of amino acids
linked by peptide bonds. These are synthesized in the granular endoplasmic
reticulum of glandular cells in the same manner as other proteins.
• The amino acid sequence is determined by specific deoxyribonucleic acid
(DNA) molecules through the messenger ribonucleic acid (mRNA)
• First the precursor hormone (prohormone, or preprohormone) is synthesized
in the rough endoplasmic reticulum.
• Then the precursor hormone is converted into proper hormone within the
Golgi complex by post-translational cleavage.
Storage and release
• Protein and polypeptide hormones are stored exclusively in subcellular
membrane-bound secretory granules within the cytoplasm of endocrine cell
until a release signal is received. On receiving the release signal, these
hormones are released into the blood by exocytosis.
3. Steroid hormones Synthesis.
• Steroids are hydrophobic lipid-soluble substances. These are
synthesized from cholesterol.
Storage and release
• There is little storage of steroids. Instead, different
precursors of cholesterol and intermediate compounds are
present in the cells.
• These serve as precursors and when the necessity of
hormone arises, the enzymatic action converts them into
steroids which are released in the circulation by simple
diffusion.
Hormone transport After secretion into
bloodstream
• The hormones may circulate in two forms:
Unbound form.
• Some hormones circulate as free molecule, e.g. catecholamines and
most peptide and protein hormones circulate unbound.
Bound form
• Some hormones, such as steroids, thyroid hormones and vitamin D,
circulate bound to specific globulins that are synthesized in the liver.
The binding of hormones to proteins is advantageous as it:
• Protects the hormone against clearance by the kidney,
• Slows down the rate of degradation by the liver and
• Provides circulating reserve of the hormone.
• Some hormones are carried in the blood as inactive forms with
proteins. They become active at the target site only. Only unbound
hormones pass through capillaries to produce their effects or to
degrade.
Plasma concentration
• Hormones are usually secreted into the circulation in extremely low
concentrations:
• Peptide hormone concentration is between 10−12 and 10−10 mol/L.
• Epinephrine and norepinephrine concentrations are 2 × 10−10 and 13 ×
10−10 mol/L, respectively.
• Steroid and thyroid hormone concentrations are 10−9 and 10−6 mol/L,
respectively. Half-life Most hormones are metabolized rapidly after
secretion.
In general:
• Peptide hormones have short half-life.
• Steroids and thyroid hormones have significantly longer half-life because
they are bound to plasma proteins.
Mechanisms of hormone disposal
The circulating hormones are disposed off by the following
mechanisms:
• Target cell uptake and intracellular degradation,
• Metabolic degradation inactivation and
• Urinary or biliary secretion.
1. Target cell uptake and intracellular degradation.
• The interaction of hormones with their target cells is
followed by intracellular degradation.
• Degradation of protein and amine hormones occurs
after binding to membrane receptors, and then
internalization of hormone receptor complex.
• Degradation of thyroid and steroid hormones occurs
after binding the hormone receptor complex to the
chromatin.
2. Metabolic degradation/inactivation.
• Only a small fraction of the circulating hormone is removed
by target tissue cells; most of the hormone extraction and
degradation occurs in the liver and kidneys.
• Metabolic degradation occurs by enzymatic processes that
include proteolysis, oxidation, reduction, hydroxylation,
decarboxylation and methylation.
• Virtually all the hormones are extracted from the plasma and
degraded to some extent by the liver.
• In addition, glucuronization and sulfaction of hormones or
their metabolites may be carried out, and the conjugates are
subsequently excreted in the bile or urine.
• 3. Urinary and biliary excretion.
• Renal clearance of hormones is reduced greatly by protein
binding in the plasma, e.g. less than 10% of secreted cortisol
appears unchanged in the urine, because only the small, free
fraction of plasma cortisol is filtered by the glomerulus.
• On the other hand, about 30% of cortisol metabolites are
excreted in urine, because they are generally unbound or
only loosely bound to protein.
• Peptide and smaller protein hormones are filtered to some
degree by the glomerulus. However, they may subsequently
undergo tubular reabsorption and degradation within the
kidney, so that only a small fraction appears in the final
urine.
Regulation of hormone secretion
• The quantity of hormones secreted is regulated in
accordance with their requirement. General
mechanisms that govern the secretion of hormone
include:
• Feedback control,
• Neural control and
• Chronotropic control.
•
1. Feedback control
• Regulation of a hormone in terms of requirement is
best accomplished through feedback from the blood
concentration of the hormone concerned (hormone–
hormone) or some result of action of the hormone
(substrate–hormone).
• Feedback control is of two types:
• Negative feedback control and
• Positive feedback control.
Negative feedback control
• Generally, the influence of blood concentration of the
hormone concerned or its effect is to inhibit further
secretion of the hormone and is called negative
feedback control
Positive feedback control.
• This is less common and acts to amplify the initial
biological effects of the hormone .
Hormonal regulation by feedback control mechanism: A,
negative feedback; and B, positive feedback.
• Depending upon the product involved, the feedback mechanism may
be:
• Hormone–hormone feedback,
• Substrate–hormone feedback and
i. Hormone–hormone feedback control.
• The best example of hormone–hormone negative feedback control is
regulation of hormone secretions by hypothalamus and pituitary, which
involves three loops :
• Long-loop feedback .
• The peripheral gland hormone (e.g. thyroid, adrenocortical and gonads)
can exert long-loop negative feedback control on both the hypothalamus
and the anterior lobe of pituitary.
• Short-loop feedback . The pituitary trophic hormones decrease the
secretion of hypophysiotrophic hormone (e.g. GHRH, GHIH, TRH, GnRH,
etc.) by short-loop feedback.
• Ultrashort-loop feedback. The hypophysiotrophic hormones may inhibit
their own synthesis and secretion via an ultrashort-loop feedback
mechanism.
Hormone–hormone negative feedback control by hypothalamus and
pituitary: A, long loop feedback; B, short loop feedback; and C,
ultrashort loop feedback.
• ii. Substrate–hormone feedback control.
• The best example of substrate–hormone feedback control is
regulation of insulin secretion from pancreatic beta cells of islets of
Langerhans and glucagon secretion from alpha cells by blood glucose
levels.
• A rise in blood glucose level promotes the secretion of insulin, while a
fall in blood glucose promotes secretion of glucagon. These responses
keep the blood glucose level within narrow limits in spite of variation
in carbohydrate intake in diet.
2. Neural control
• Neural control acts to evoke or suppress hormone secretion in response
to both external and internal stimuli.
• External stimuli which can modulate hormone release through neural
mechanisms may be visual, auditory, olfactory, gustatory and tactile.
• Internal stimuli which influence hormonal release through neural
mechanism include pain, emotion, sexual excitement, fright, stress and
changes in blood volume.
• Neural control depending upon the type of nerve fibres involved may
be:
• Adrenergic,
• Cholinergic,
• Dopaminergic,
• Serotoninergic and
• Gabaergic.
• Examples of neural control of hormones are:
• Release of oxytocin, which fills the milk ducts in response to the
stimulus of suckling,
• Release of aldosterone, which augments the circulatory volume in
response to upright posture and
• Release of melatonin in response to darkness.
3. Chronotropic control
• Chronotropic control of hormone secretion accounts for:
Oscillating and pulsatile release of certain hormones,
• Diurnal variation in hormonal levels,
• Menstrual rhythm,
• Seasonal rhythm and
• Developmental rhythm.
• The source of regular oscillatory cycles is a pulse generator(s) located
in the suprachiasmatic nucleus (SCN) of the hypothalamus
Hormone: Receptors and mechanism of action
Hormone receptors
• All hormones act through specific receptors. Almost all hormone
receptors are large proteins present in hormone-sensitive target cells.
Characteristics of hormone receptors
• Receptor specificity
Receptor location.
• Depending upon the location, receptors are of two types:
• i. Internal receptors are located inside the cells, e.g. receptors for steroid
hormones and thyroid hormones are localized within the nucleus of
target cells.
• ii. External receptors are located on the plasma membrane of target
cells, e.g. peptide and protein hormones, amines and prostaglandins are
interspersed within the phospholipid bilayer of the plasma membranes.
• Receptor affinity refers to the degree of attraction between the
hormone and receptor and is measured from the speed of hormone
binding by the receptor. Receptor capacity refers to the quantity of
hormone binding.
• Receptor density
• Approximately 104–105 receptors exist on the surface of a polypeptide
hormone target cell, and about 3 × 103–3 × 104 intracellular receptors
exist per steroid target cell.
Change in receptor number
• Number of receptors of a cell vary depending upon the situation. It is
regulated by two mechanisms: downregulation and upregulation.
• i. Downregulation refers to decrease in the number of active receptors. It
occurs to regulate the hormone sensitivity when it is present in excess. For
example, elevated ambient insulin concentration causes a loss or
inactivation of insulin receptors in liver cells, fat cells and white blood cells
(WBCs).
• ii. Upregulation refers to increase in the number of active receptors on a
cell. It occurs to regulate the hormone action when its concentration is
less. This phenomenon tends to reduce the effect of hormone deficiency.
• Spare receptors. A maximum physiologic response of a target cell is
observed even when the concentration of a hormone is lower than
required to occupy all of the receptors on that cell. Therefore, most of the
receptors (∼97%) are referred to as spare (reserve) receptors.
• Classification of membrane receptors Cell membrane receptors have
been classified into four classes
Schematic general structure of four major classes
of membrane receptors
1. Receptor kinases. These include membrane receptors that
contain enzyme activity. For example, receptors for insulin
hormone contain tyrosine or serine kinase as an intrinsic part
of structure.
2. Receptor-linked kinases. These have no intrinsic enzyme
activity, e.g. receptors for growth hormone, prolactin and
cytokines.
3. G-protein-coupled receptors. Examples are: receptors for
pituitary tropic hormones, glucagon, epinephrine
norepinephrine, parathyroid hormone and prostaglandins.
4. Ligand-gated ion channels. These act as receptors for
neurotransmitters such as acetylcholine (ACh), γ-aminobutyric
acid (GABA) and glycine.
Structure of a receptor
• The receptors in general have two parts or domains: a recognition
domain (R), and a coupling domain (C)
• Recognition domain (R): It is that part of the receptor where the
hormone binds. It is formed by extracellular N-terminus portion of
the receptor (extracellularly projected NH2 group).
• Coupling domain (C): It is that part of receptor which initiates signals
for intracellular activities after the recognition. It is formed by the
intracellular C-terminus tail of the receptor (intracellularly projected
carboxyl, i.e. COOH).
• The membrane receptors wind in and out of the plasma membrane
by means of seven transmembrane segments.
Mechanism of action of hormones
• All hormones act through specific receptors, and depending upon the
mechanism of action hormones they have been divided into group I
and group II .
• Group I hormones act by affecting gene expression at cellular level,
and Group II hormones act through intermediary molecules called
second messenger, and depending upon the chemical nature of
second messenger, the group II hormones have been further divided
into subgroups A, B, C and D .
General structure of a hormonal receptor
The main mechanisms of hormone actions
are:
• Action through change in membrane permeability,
• Action through effect on gene expression by binding of hormones
with intracellular receptors,
• Action through secondary messengers which activate intracellular
enzymes when hormones combine with membrane receptors and
• Action through tyrosine kinase activation.
Action through change in membrane
permeability
• Some hormones bind with the receptors present in the cell
membrane (external receptors) and cause conformational change in
the protein of the receptors, this results into either opening or closing
of the ions channels (such as Na+ channels, K+ channels and Ca2+
channels).
• The movement of ions through Ca2+ channels causes the subsequent
effect, e.g. adrenaline, noradrenaline act by this mechanism.
Action through effect on gene
• Action through effect on gene expression by binding of hormones with
intracellular receptors Group I hormones act by their effect on gene
expression which include steroid hormones, retinoids and thyroid hormones.
• These hormones are lipophilic in nature and can easily pass across the cell
membrane.
• They act through intracellular receptors located either in the cytosol or in the
nucleus.
The sequence of events involved is
• 1. Transport. After secretion, the hormone is carried to the target tissue on
serum-binding protein.
• 2. Internalization. Being lipophilic, the hormone easily diffuses through the
plasma membrane.
• 3. Receptor–hormone complex is formed by binding of hormone to the
specific receptor inside the cell.
• There are three distinct pathways by which primary messenger alter
transcription and gene expression as under:
• Primary messenger (in case of steroid and thyroid hormone) directly
binds to nuclear receptor and directly interacts with DNA to alter
gene expression.
• By activation of cytoplasmic receptor protein (kinase) that moves into
the nucleus and activates latent transcription factor causing
phosphorylation, e.g. mitogen-activated protein kinase (MAP).
• The third pathway is the activation of latent transcription factor
present in the cytosol. This pathway is shared by nuclear factor kappa
B (NFKB) and signal transducers of activated transcription (STATs).
• 4. Conformational change occurs in receptor proteins leading to
activation of receptors.
• 5. The activated receptor–hormone complex then diffuses into the
nucleus and binds on the specific region on the DNA known as
hormone responsive element (HRE), which initiates gene
transcription. 6. Binding of the receptor–hormone complex to DNA
alters the rate of transcription of messenger RNA (mRNA).
• 7. The mRNA diffuses in the cytoplasm, where it promotes the
translation process at the ribosomes. In this way, new proteins are
formed which result in specific responses. Some of the new proteins
synthesized are enzymes.
Action of hormones through their effect of gene
expression
Action through second messengers
• The peptides and biogenic amines are two principal classes of
hormones which act through second messengers and are classified as
group II hormones . Such hormones are also called first messengers.
The release of second messenger is mediated by GTP-binding proteins
also called G-proteins Coupling by G-proteins.
• The G-proteins involved in cell signalling are divided into two groups:
small G-proteins and heterotrimeric G-proteins.
(i) The small G-proteins further belong to six different families or small
GTPases and act as follows:
• The GTPases cause hydrolysis of GTP to GDP and tend to inactivate
small G-proteins.
• Guanine exchange factors (GEFs), present on the active site, activate
G-protein by encouraging exchange of GDP for GTP.
• Some small G-proteins lead to lipid modification and thus help to
anchor them to the membrane.
• Some small G-proteins are free to diffuse into the cytosol.
Functions:
• The Rab family members of small G-proteins regulate vesicular
movement between endoplasmic reticulum, the Golgi apparatus,
lysosomes and cell membrane.
• The Rho/Rac family small G-proteins mediate interaction between
cytoskeleton and cell membrane.
• Ras family small G-proteins are responsible for regulation of growth
by transmission of signals from the cell membrane to the nucleus.
• (ii) Heterotrimeric G-proteins are large G-proteins that couple the
receptors to catalytic units on the cell surface and catalyse the formation of
intracellular second messenger.
• The heterotrimeric G-proteins are made up of three subunits (α, β and γ).
The α and γ subunits help in anchoring of G-protein to the cell surface . The
α subunit is bound to GDP.
• When hormone or ligand binds to G-protein-coupled receptor the GDP is
exchanged with GTP and α subunit separates from the β and γ subunits and
is responsible for various biological activities. The intrinsic GTPase activity
of separated α subunit converts GTP into GDP and leads to reassociation of
α subunit with β and γ subunits.
• The regulators of G-protein signalling (RGS) accelerate α subunit GTPase
activity.
• The heterotrimeric G-protein involved in relaying signals for more than
1000 G-protein receptors
• G-protein-coupled receptors structurally (GPCRs) are barrel like.
• The G-protein-coupled receptors (GPCR) are seven helix or serpentine
receptors and span the cell membrane seven times.
• Mechanism of action:
• When a ligand binds the GPCR, a conformational change occurs,
which activates heterotrimeteric G-protein-associated cytoplasm, i.e.
leaf of plasma membrane which in turn activates many G- proteins,
thus leading to amplification of the effect.
• A large number of ligands act through G-protein-coupled proteins.
These include: .
• Events involved in coupling by G-protein which lead onto changes in
the cellular concentration of the second messengers are summarized
• Group II hormones are water soluble and bind to the plasma
membrane of the target cell via cell surface receptors.
• - The hormone-bearing receptor then interacts with a G- protein and
activates it by binding GTP. There are two classes of G-proteins:
stimulatory G-protein (Gs) and inhibitory G-protein (Gi).
• - In its activated (“on”) state, the G-protein interacts with one or more
of the effector protein (most of which are enzymes or ion channels
such as adenylyl cyclase, Ca2+ or K+ channels or phospholipase C, A2
or D) to activate or inhibit them.
• - The changed effector molecules in turn generate second messenger
that mediates the hormone’s intracellular action.
Schematic mechanism of coupling by G- protein leading to
increase in second messenger which mediates hormone’s
physiological response.
• The second messenger systems that are activated through coupling of
hormone receptor complexes by G-protein include
• Adenylyl cyclase–cAMP system,
• Guanylyl cyclase–cGMP system,
• Membrane phospholipase–phospholipid system and
• Calcium–calmodulin system.
1. Adenylyl cyclase–cAMP system
• The adenylyl cyclase–cAMP system was the first to be described by
Sutherland in 1961 that initiated the concept of second messenger. The
hormones which act through this system constitute the group IIA
hormones . The steps involved in the hormone action via adenylyl cyclase–
cAMP system are summarized below :
• i. Binding of hormone (Step 1) to a specific receptor in the cell membrane.
• ii. Activation of G-protein (Step 2). After formation of hormone–receptor
complex, the GDP is released from the G-protein and is replaced by GTP,
i.e. G-protein is activated.
• iii. Activation of enzyme adenylyl cyclase (Step 3).
The hormone– receptor complex via activated G-protein (stimulatory or
inhibitory) either stimulates or inhibits the enzyme adenylyl cyclase which is
also located in the plasma membrane.
• iv. Formation of cAMP (Step 4).
A part of the enzyme adenylyl cyclase protrudes through the inner
surface of the cell membrane and when activated it catalyses the
formation of cAMP from cytoplasmic ATP with Mg2+ as cofactor. A
stimulatory G-protein (Gs) therefore increases intracellular cAMP
levels, whereas an inhibitory G-protein (Gi) decreases cAMP levels.
v. Action of cAMP.
The cAMP once formed stimulates a cascade of enzyme activation. One
molecule of cAMP may stimulate many enzymes. Therefore, even the
slightest amount of hormone acting on the cell surface can initiate a
very powerful response.
The cyclic AMP so formed initiates response by different mechanisms in
eukaryotic and prokaryotic cells.
• In eukaryotic cells, the cAMP activates protein kinase A (Step 5) which
phosphorylates the specific proteins, producing highly specific physiological
actions (Step 6):
• Increase in cell permeability (e.g. ion transport and secretion),
• Increase in synthesis of enzyme system (enzyme induction),
• Activation of enzymes present in the cell.
• Release of hormones from storage pack cells and steroidogenesis,
• Gene regulation
• In prokaryotic cells, in contrast to eukaryotic cells, the cAMP acts directly at
the gene level. It binds to a specific protein called catabolic regulatory protein
(CRP). In association with CRP, it binds to DNA and influences gene expression
or transcription. Therefore, cAMP in prokaryotic cells acts just like steroid
hormones.
• vi. Inactivation of cAMP. The cAMP is degraded to 5’-AMP (inactive form) by
enzyme phosphodiesterase. Hence, the effect of cAMP is short lived if the
hormone stimulating adenylyl cyclase is removed. Phosphodiesterase
inhibitors such as caffeine and theophylline would be expected to augment
the physiologic action of cAMP.
Mechanism of action of hormone through adenylyl cyclase
(cAMP) system as second messenger.
2. Guanylate cyclase–cGMP system.
• Group II-B hormones which act via second messenger cGMP include
atrial natriuretic factor (ANF) and nitric oxide (NO).
• i. Synthesis of cyclic GMP is analogous to the formation of cAMP.
Enzyme guanylate cyclase produces cGMP from GTP.
• ii. cGMP exerts its biochemical response through an enzyme protein
kinase G, which when activated initiates a cascade of subsequent
enzyme activations that is characteristic of this signalling system.
• Note: Cyclic GMP is important in vision. Both rod and cone cells
contain ion channels which are regulated by cGMP.
3. Membrane phospholipase–phospholipid system or
inositol triphosphate IP3 mechanism
• Hormones which exert their response through this system constitute
the so-called group II-C hormones .
• Steps involved in this system are :
• Hormone binds to a receptor in the plasma membrane.
• The hormone–receptor complex via a G-protein activates the
membrane enzyme phospholipase C present on the inner surface of
the membrane.
• Activated phospholipase C (PLC) then releases diacylglycerol and
inositol triphosphate (IP3) from membrane phospholipid.
• Inositol triphosphate (1,4,5 triphosphate) or (IP3) then mobilizes Ca2+
from the endoplasmic reticulum.
• Calcium ions (Ca2+) and diacylglycerol together activate protein kinase
C.
• Activated protein kinase C phosphorylates proteins and causes
specific physiological action.
• Diacylglycerol also yields arachidonic acid which serves as a substrate
for rapid synthesis of prostaglandins that modulate cell response.
Mechanism of action of hormone via membrane
phospholipase–phospholipid system or IP3 mechanism.
4. Calcium–calmodulin system.
• Hormones that act through this system as second messenger are also
included in the so-called group-II C hormones .
• Steps involved in this system are :
• Hormone binds to a specific receptor in the plasma membrane.
• Then the hormone–receptor complex, via G-protein opens the Ca2+
channels on the cell membrane and also activates mobilization of
Ca2+bound to the endoplasmic reticulum.
• Ca2+ binds to a specific binding protein, calmodulin, in various
proportions.
• The different calcium–calmodulin complexes activate or deactivate
various calcium-dependent enzymes producing different physiologic
actions.
Mechanism of action of hormone via
calcium– calmodulin system.
Mechanism of action of hormone via tyrosine
kinase activation
• Certain hormones act by activating tyrosine kinase system
and have been classified as group-II D hormones .
• This mechanism of signal generation from plasma membrane
receptors does not require G-protein intermediaries.
• These receptors have an extracellular hormone-binding
portion, a single transmembrane portion and an
intracytoplasmic C- terminal portion.
• The activation of tyrosine kinase occurs by two mechanisms:
1. Hormone receptors possessing intrinsic tyrosine activity, e.g. those
for insulin and epidermal growth factor (EGF) involve the following
steps :
• Binding of hormone to the receptor changes its conformation and
exposes sites on its intracellular portion that are capable of receptor
autophosphorylation at specific tyrosine sites.
• As a result, the receptor itself becomes a tyrosine kinase that
phosphorylates tyrosine residue on intracellular protein substrates.
• This latter activity sets into motion a cascade of events leading to
enzyme activation and gene transcription.
2. Hormone receptors that do not possess intrinsic tyrosine activity, e.g.
those for growth hormone, prolactin-releasing hormones, cytokines etc. act
by activation of JAK2–STAT pathway.
• JAK2 belongs to Janus family of tyrosine kinases. The phosphorylation of
tyrosine kinases causes activation of STATs (signal transducer and activators
of transcription) which are cytoplasmic transcription factors.
• On activation, STATs migrate into the nucleus and causes transcription of
various genes as follows :
• Hormone binding to extracellular portion of the receptor changes its
intracytoplasmic tail.
• The changes produced in intracytoplasmic tail of receptor expose sites
which attract and dock the intracytoplasmic tyrosine kinases (such as JAK
kinases and STAT kinases) and then activates them.
• The activated intracytoplasmic tyrosine kinases phosphorylate cytoplasmic
substrates such as transcription factor proteins and ultimately modulate
gene expression.
Mechanism of action of hormone via tyrosine kinase activity: A, by
receptors that possess intrinsic tyrosine activity; and B, by receptors that act
by activation of JAK2– STATs pathway.
 Measurement of hormones (READ UP)
• Measurement of blood level of hormones is essential to confirm the
endocrinal disorders associated with either deficiency or excess of a
hormone.
• Since hormones exist in the blood at very low concentrations, the
conventional methods of estimation, such as colorimetry, are not of much
use.
• Therefore, they are measured by hormone assays and some special
techniques which include:
• Bioassay,
• Immunoassay,
• Cytochemical assay and
• Dynamic tests.
• Note. The hormonal action mediated through intracellular receptors
is comparatively slower due to the time (45 min to several hours)
involved in the processes described above. Therefore, glucocorticoids
may take hours to few days to achieve the therapeutic effect.
Bioassay Biological assay
• Bioassay Biological assay that was frequently used earlier to measure
the level of a hormone in the plasma is now an obsolete technique.
• In this method, hormone levels were assessed by injecting the
unknown sample of plasma in experimental animals and observing
quantitatively the specific biological effect.
• The effect chosen was a characteristic action of the hormone for
which a clear dose–response relationship existed. For example, one
unit of insulin was defined as one third of the amount of insulin that
will lower the blood sugar of a rabbit weighing 2 kg to conclusive
levels in 3 h
Immunoassay
• The immunoassay methods, frequently employed for estimation of hormone
levels, include:
• Radio immunoassay (RIA) and
• Enzyme-linked immunosorbent assay (ELISA).
• 1. Radioimmunoassay The radioimmunoassay (RIA) was developed by Bershon
and Yalow in 1960s, for which they were awarded the Nobel Prize in late
1970s. This test is performed as:
• An unknown sample of plasma in which the concentration of a particular
hormone (H) to be estimated is mixed with commercially available purified
specific antibody (anti-H) and an appropriate amount of the purified hormone
tagged with radioactive isotope (H+). The mixture is incubated in cold.
• The antibodies have high affinity for the hormone. There occurs a competition
between the free hormone (H) present in the unknown sample of plasma and
the tagged hormone (H+) for binding to the specific antibody (anti-H).
2. Enzyme-linked immunosorbent method
Enzyme-linked
• immunosorbent (ELISA) method is principally similar to RIA, i.e. it is
also based on the principle of antigen–antibody reaction.
• Any antigen that is protein can be measured by this technique. In this
method, radioactivity is not measured; instead specific antibody
hormone (antigen) complex is stained with suitable dye, e.g. di-
ammonium 2-2’ azinobis (3-ethylbenzothiazolene 6 sulphate), also
called ABTS, and the intensity of colour is measured by
spectrophotometer. This technique is useful in estimating peptide and
steroid hormones.
Cytochemical assay
• This test is much more sensitive than the immunoassay, but
is cumbersome and time consuming and so rarely used.
• In this technique, genesis of hormone can be detected in
slices cut out of the endocrine gland by incubating them in
an ascorbate-enriched culture medium.
• This test is very useful in measuring the minute basal levels
of hormone secretion.
Dynamic tests
• Dynamic tests are needed in certain situations when simple blood
hormone level estimation is not enough. Two types of dynamic tests
are: Suppression type of dynamic tests are useful in certain
conditions, e.g. to know whether a lung cancer is secreting ACTH.
Stimulation type of dynamic tests are useful in certain other
conditions, e.g. metyrapone test is performed to know whether the
corticotrophs of the pituitary (which secrete ACTH) are normally
functioning or not.