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pharm
2/24/98

fluharty

we're still discussing autacoids - 
compounds of diverse chemical nature involved in local tissue responses. 
we spoke before about the renin/AT system,which is a classic hormonal 
system involving cardiovascular homeostasis, but also has paracrine 
mechanisms.

we're going to talk about bradykinin, serotonin,and 
histamine today.

the histamine lecture is new. 

bradykinin:
this system 
is related to the renin AT system partly via physiological actions,and 
partly b/c they share an enzyme. but the bradykinin system is more 
complex, though similar.

bradykinin story starts in 1920s with discovery 
of an enzyme. kallikrein was found (there are really multiple forms of 
this, tissue and plasma forms), it was characterized as a hypotensive 
substance in urine, saliva, plasma, and tissues. (opposite of renin). by 
40s-50s, they realized this enzyme was part of a larger system that 
ultimately generated bioactive things like bradykinin and kallidin. 
kallidin is mostly in tissue, bradykinin in plasma and some in tissue.


now, there are many drugs that antagonize actions of kallidin and 
bradykinin at their receptors. many of these antagonists are peptides, 
based on the structures of the peptides. usually,though, peptide 
antagonists aren't good to be given systemically - work better used 
locally.

we now know there are two peptides making up the overall 
bradykinin system - kallidin and bradykinin (kallidin has an extra AA on 
it). 

there are a variety of inflammatory events that activate this 
system. the events that turn this on are more complex than those of the 
renin/AT system - it involves collagen, clotting, local inflammatory 
events, stress to tissues. also, it's more complex because it has two of 
everything. two precursors - high mw kininogen and low mw kininogen. two 
enzymes - tissue and plasma forms of kallikrein.
and two (at least) 
biologically active peptides - bradykinin and kallidin - and two 
degradative enzymes - kininase I andII

precursors: these are substrates 
acted on by the enzymes. they are large plasma alpha globulins 
synthesized in the liver, like angiotensinogen. the high mw form is a 
substrate for both enzymes - both plasma and tissue kallikrein can 
generate peptide from the high mw precursor. the low mw form results in 
substrate with smaller carboxy terminus, due to alternate mRNA processing 
of the same gene used for the high mw form. this form is really only used 
to make kallidin, can't be used to make bradykinin. so the two substrates 
are different in size, and one can yield either bioactive peptide, one 
can only yield kallidin.

enzymes: associated with plasma or tissue. 
unlike renin and ACE, these are largely inactive. they need to be 
activated =- the way they are activated is by specific proteases cleaving 
off inhibitory enzymes, allowing them to act. this is somewhat true for 
prorenin/renin but we didn't go into that. these enzymes are inactive and 
must be activated by proteases - and therefore protease inhibitors can 
actually decrease activity in this system, by preventing liberation of 
active enzymes. 

so you end up with bradykinin and kallidin which are 
interchangeable at a tissue level, because if you cleave off part of 
kallidin, you end up with bradykinin. like all peptides,once these are 
made they have very short half lives - one pass through lungs destroys 
90% of it. degradation is via kininase I and II - kininase II is 
identical to ACE (angiotensin converting enzyme) - this is the link b/w 
these systems. remember, ACE generates ATII - here, ACE, which we now 
call kininase II, breaks down/inactivates bradykinin. 

you may recall we 
used ACE inhibitors to block renin/AT system - side effects included skin 
rashes, cough - it's argued that these effects are due to potentiating 
the effects of bradykinin. that's undesirable - the drugs we use are 
blocking this rather nonspecific enzyme. 

in addition, there is also a 
more unique enzyme, kininase I, and it has been argued that this enzyme 
generates fragments that stay active in some damaged tissues, b/c tissue 
damage induces a receptor for these fragments. it's not clear that 
kininase I is truly degradative as much as it is a local enzyme 
generating another locally biologically active substance.

what does this 
system do? 

bradykinin and kallidin are implicated in many inflammatory 
events - this includes inhalation of AG that produce pulmonary irritation 
and edema, endotoxic shock, hereditary angioedema - an episodic 
swelling/edmea syndrome associated with excessive acctivation of plasma 
kallikrein, lots of inflammatory events are involved. we'll focus on some 
of the pulmonary events which are most important pharmacologically. 

bradykinin receptors are seen throughout the nervous system, and are 
associated with nociception. also there's evidence that kinin systems in 
kidney play a role in renal function - bradykinin can produce increased 
Cl- transport,may act in LOH to promote volume expansion. these renal 
effects could start to augment the renin/AT system as well.

also, 
bradykinin/kallidin may act to lower BP.
pharmacologically, these are 
potent vasodilators because bradykinin receptors on endothelial cells 
produce increased Ca++ levels, and so are coupled to the formation of 
endothelium derived relaxing factor, aka NO, Time Magazine's former 
Molecule of the Year :) NO then diffuses from endothelial cell to smooth 
muscle cells, activates cGMP, and produces vascular relaxation. 


serotonin and histamine share this same NO-inducing property, and also 
cause vasodilation.

bradykinin may produce reflexive tachycardia as a 
consequence of lowering BP via vasodilation. also increases vascular 
permeability and this underlies formation of edema

what goes on here, 
it's kind of a disruption of starling equilibrium. remember, on arterial 
side of capillary beds,  hydrostatic forces overwhelm plasma protein 
forces, tissue exits - and then on venous side, hydrostatic pressure is 
lower, proteins pull fluid back into vessels. if you increase vascular 
permeability, proteins leak out, and then fluid won't come back into 
vessels on venous side. fluid gets trapped in tissue, producing edema. 
this is true with all these autocoids.

in most nonvascular smooth 
muslce, bradykinin produces contraction - causes bronchoconstriction - 
this is one reason why there are efforts to make antagonists for use in 
allergic airway disease.also this is why ACE inhibitors may cause 
coughing - bradykinin induced coughing and pulmonary irritation

also 
causes slow burning pain at nerve endings.

there are at least three 
described bradykinin receptors - B1 is most important in contraction of 
some  vascular beds, and is sensitive to product of kininase I acting on 
bradykinin. this receptor may be induced by tissue damage. 
B2 receptors 
mediate almost all the other things associated with kinin system - except 
that the B3 receptor is the one in tracheal smooth muscle, and is 
involved in bronchoconstriction - this receptor is targeted by developing 
drugs for use in allergic airway syndromes or for use along with ACE 
inhibitors. these are all Gprotein coupled receptors, usually involving 
phospholipase C and a rise in Ca++ producing a rise in NO. 

drugs:
most 
have been somewhat limited since they're peptides. these are usually only 
good for local application b/c they are so rapidly degraded by one pass 
through lungs.

new B3 antagonists have had some usefulness treating cold 
viruses, asthma, or skin pain. 

another approach is to target the 
enzymes which are normally inactive, and whch must have inhibitory 
sections removed by proteases. if you block the proteases, you reduce the 
overal enzymatic activity of the system. there are drugs that dothis - 
aprotinin (??) used in acute pancreatitis. other compounds are being 
developed that attack other cleavage sites.

like renin/AT system, 
there's reason to think that many drugs will exploit this system and 
local regulation of inflammation, edema, vasodilation, and 
bronchoconstriction. 

bradykinin and renin/AT systems are two classic 
peptide systems involved in autocoid capacity.

shifting gears - two very 
different kinds of chemical compounds: biogenic amines. serotonin and 
histamine.

serotonin:

recall that this is a biogenic amine - like all 
of them,it has a complex cyclic carbon structure, here a hydroxylated 
indol ring, a two carbon separation, and a terminal amine. we know that 
serotonin is very important as a neurotransmitter - it's distributed 
throughout the brain, the raphe nuclei, distributed into the cord, etc. 
but today we're interested in its autacoid functions - vascular control, 
edema formation, GI function, etc. 

serotonin is so important it was 
discovered twice! here, it was discovered as a vasoconstricting substance 
associated with clotting - we called it vasotonin. it's abundant in 
platelets and is released during clotting activity. also causes 
vasoconstriction at that time. in 1959 it was synthesized and seen to be  
5-hydroxytryptamine. at the same time, a gut factor called enteramine was 
isolated from the gut.most of the serotonin in your body is in 
enterochromaffin cells, and is released in response to pH changes, 
osmotic changes, food, anything. well, this enteramine was also isolated 
as 5-hydroxytryptamine.
so serotonin is abundant in platelets, gut,and 
really everywhere. it's in every animal on the planet,in fruits, nuts, 
mushrooms, venoms, - local depositing of serotinin when you're stung by a 
bee is in part responsible for edema.

in mammals, 90% of it is in 
enterochromaffin cells, the rest in platelets, CNS, and mast cells 
(although not much serotonin in human mast cells - they have more 
histamine).

serotonin and histamine share many qualities.

anyway, in 
all cells in body (except platelets which get their serotonin by 
scavenging it generally from the gut - platelets don't have the enzymes 
to make serotonin. this is key), serotonin is synthesized as per figure 2 
in the handout. you start with tryptophan - all cells which make 
serotonin have high affinity tryptophan uptake systems. tryptophan 
hydroxylase is the ratelimiting enzyme in serotonin synthesis. it 
converts tryptophan into 5hydroxytryptophan.then an AA decarboxylase 
takes off the carboxy group, creating 5HT aka serotonin. decarboxylation 
is critical in generation of all biogenic amines. you always 
decarboxylate your AA precursors to make active biogenic amines. then, 
serotonin is stored in vesicles, and released on stimulation of the cell 
it's in. it's degraded mainly by MAO, followed by oxidation/reduction and 
excretion in urine. when serotonin functions as autocoid, action can also 
be terminated by platelet reuptake of serotonin as well - especially in 
gut. 

actions of serotonin:

acts in abundant fashion in the brain - 
regulation of sleep, cycling,pain perception and control, autonomic 
function, affective disorders including unipolar depression in humans. 
it's now also known to be important regulator of food intake - both 
number and frequency of meals. also has prominent neuroendocrine effects 
- involved in control of all hormones released from anterior pituitary - 
involved in hypothalamic hypophyseal portal system. also may regulate 
release of thyroxin, insulin. is a precursor for melatonin in the pineal 
gland as well. 

in GI system, serotonin is an important autacoid. it's 
richly present in enterochromaffin cells, very important in regulation of 
GI function. it's colocalized with substance P - which might also be 
considered an autacoid, but is best defined as a mediator of pain in the 
nervous systme. serotonin is released by mechanical stimulation 
associated with food, hypertonicity, osmotic challenges - all food we eat 
is of greater osmolarity than plasma, so it always signals GI tract to 
regulate food transit. gastric acid secretion, norepi, and vagal tone - 
basically everything that changes when you eat a meal, results in release 
of serotonin, which we know helps regulate GI motility, facilitating 
movement of food through GI tract.

adding on top of this the realization 
that serotonin in brain is involved in appetite, you get an interesting 
picture of desire to eat, frequency of eating, appetite, and metabolic GI 
function. this is why we're trying to use drugs to affect serotonin 
levels as antiobesity agents.

serotonin effects on platelet function - 
it's abundant in platelets, but we're not really sure what the function 
is. bleeding time is largely unaffected by serotonin dpeletion. it may 
have something to do with vascular permeability, but we're not sure of 
the function.

pharmacologically-

high levels of serotonin that develop 
secondary to drug administration produce certain effects - that turned 
out to be a problem with fenfluramine - including bronchoconstriction, 
complications of a pulmonary nature. in cardiovascular system, one key 
thing is positive inotropic and chronotropic actions on heart, and maybe 
toxicity to the heart develops with excessive serotonin release.
in 
smooth muscle - promotes GI motility in lowdoses.
in nervous system, high 
levels of serotonin produce release of catecholamines from adrenal 
medulla, adding to potential cardiac toxicity. 

how does this all work? 

mostly receptor mediated. it interacts w/a variety of receptors. the 
advent of molecular biology has given us an embarrassment of riches.in 
past 15 yrs, more and more 5HT receptors have been discovered. there are 
families known as 5HT1 like, 5HT2 like, 5HT3 like, 5HT4, 5HT5, 5HT7, and 
really up to 25 different receptors have been described. so our handout 
is already outdated. this is only important in realizing that the really 
good drugs are highly specific for receptors. receptor diversity is 
nature's invitation for developing highly specific drugs.

the 5HT3 
receptor is exception to rule. most of what serotonin does is mediated by 
second messengers - gprotein coupled receptors. but 5HT3 recep is ligand 
gated ion channel promoting cation influx and fast depolarization. 


drugs affecting serotinergic systems:

1. those that affect endogenous 
serotonin levels - we tried precursor loading with tryptophan - giving 
more tryptophan to increase serotonin levels - may be useful in 
phenylketonurics. also lots of popular press suggestions that this 
improves sleep. you can no longer buy tryptophan in health stores. 
inhibitors of synthesis to try to shut down serotonin production have 
almost always been toxic. clinical uses aren't around.

2. uptake 
inhibitors - serotonin is inactivated by high affinity uptake. inhibitors 
block uptake by platelets, neurons, etc. fluoxetine aka prozac is the key 
ssri - commonly prescribed. fenfluramine is a complex serotonergic 
compound that can interfere with uptake, promote serotonin release, and 
stimulate 5HT2 receptors in the brain - totally jacks up serotonin 
system! this is why it worked so well with appetite regulation - but had 
lots of bad effects too.

3. receptor antagonists - many types. most of 
them are mixed compounds. we don't have good selective compounds. most 
are affecting several types. methesergide (? similar to LSD w/o 
hallucination effects) used for migraines, diarrhea treatment in patients 
with enterochromaffin tumors. 

other compounds that are serotinergic and 
histaminergic.
other sertoinergic compounds are touted for vascular 
control -but this is probably due to alpha receptor interaction.

the big 
drugs are the 5HT3 antagonists - the ones Dr Washabau talked about. these 
are the ones that are able to work miracles in blocking nausea. general 
anesthesia causes horrible nausea. odansitron (?) is given when you're 
put under, now, and it totally eliminates the nausea. it's also great for 
use with chemotherapy. these work through a series of reflexes involving 
the CRTZ (AP), the ability to respond to circulating toxins, and various 
medullary vomiting reflexes.

other serotinergic antagonists in CNS will 
probably become important in the future. and of course the antagonists 
are important for use in patients with enterochromaffin tumors. 

really, 
you could look at serotonin assomething to control food intake, appetite, 
regulate metabolism, and when you go to far, regulating nausea and 
vomiting. 
 
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histamine

this is the last autacoid we have to 
discuss. it's also a biogenic amine like serotonin and has similar 
structural properties - the classic here is an imidazole ring separated 
by two carbons from the terminal amine. production involves 
decarboxylation of histidine.

isolated in 1927 from lung and liver. now 
we know it is found in tissues all over almost all animals and plants. 
concentrations vary dramatically from tissue to tissue and species to 
species. literally, the term histamine comes from histos which means 
tissue - because it's everywhere. "tissue amine" 

it's generally thoguht 
that most histamine is associated with mast cells, and this is the 
predominant site in most animals and in humans.  skin, bronchi, GI mucosa 
- tons of histamine. not always in mast cells, must most predominant in 
tissues with lots of mast cells.

is released via many mechanisms, not 
unlike bradykinin system - injury, trauma, burn, allergies all promote 
histamine release. surface acting compounds, irritants, etc. many 
chemicals, some of which you use in other contexts,like morphine, curare, 
etc can also promote histamine release. sometimes this is bad.


histdidine ->histidine decarboxylase acts on it to make histamine by 
removing a carboxyl group. this is the rate limiting step. drugs blocking 
histamine synthesis selectively inhibit histidine decarboxylase. in many 
tissues,histamine turnover is very slow. if you block synthesis, can have 
low levels for weeks. histamine is broken down by two enzymes - most 
commonly by a methylating enzyme, called histamine-n-methyl transferase, 
followed by oxidative deamination and urinary excretion. or can be broken 
down by diamine oxidase aka histaminase - but this is less common.


histamine does all it's things that it does by interacting with three 
well recognized classes of receptors - see table in handout. there are 
H1, H2, and H3 receptors. we all probably have personal experience with 
H1 receptor antagonists - these are commonly sold OTC for allergies, etc. 
some of us may also have experience w/H2 receptor antagonists that 
control gastric acid secretion - tagamet, zantac. H3 receptors seem to be 
involved in histamine release - not really clinically targeted.

H1 is 
classical receptor known for many years. histamine's terminal amine first 
interacts with the receptor. histamine receptors are mapped throughout 
the body and are clearly very important in bronchi and in gut. effector 
pathway is stimulation of phosphoinositide metabolism. increase in Ca++ 
levels in cells, production of NO, diffusion to muscle, elevation of 
cGMP, and vasodilation. don't have to know the names of the drugs in the 
tables. the second messenger pathway ISimportant. 

H2 receptors - 
mediate stuff that H1 doesn't mediate - important for uterine motility, 
chronotropic effect on heart. things that bind to this receptor bind 
first with the imidazole portion, not the amine first like the H1 
receptor.this may be helpful for making more selective drugs in the 
future. H2 receptors are mapped throughout the body - in tracheal sm 
muscle, heart, gut. second messenger system seems to be production of 
cAMP. cAMP is important for regulation of gastric acid secretion.

H3 - 
involved in mediating rate of histamine release - in nerve endings and 
cells that store histamine. beyond that,we're not sure what these 
receptors are for. they're found in many tissues that also express H1 and 
H2, but in lower numbers. most important thing to know about them that 
may be useful later is that they may regulate synthesis and activity of 
histidine decarboxylase, and release of histamine from mast cells. 
this 
receptor may produce a decrease in cAMP production - so if H2 and H3 are 
on same cell, may antagonize eachother.

histamine has highest affinity 
for H1 receptor.this helps to figure out how H1 can initially mediate an 
effect that later is sustained by other receptor subtypes.

what are some 
effects of histamine?

no really important effect on heart - maybe 
reflexive tachycardia due to decreased BP but this is trivial.usually, 
prominent effect is vasodilation, but there is some contraction in some 
vascular beds, usually mediated by H1 receptors. in capillaries, the big 
effect is marked dilation - balance of H2 and H1 receptors is important 
here - you get rapid, large, transient dilation from H1 stimulation =- H2 
stimulation gives sustained,long lasting dilation - due to 2nd 
messengers, most likely - the cAMP elevation is more long lasting. also, 
as with autacoids, there are changes in permeability mediated by H1 
receptors via increased Ca++ levels, resulting in plasma protein leakage, 
interstitial fluid expansion and edema. this is standard autacoid 
mechanism - disruption of starling equilibrium allowing proteins to flow 
out, trapping fluid in interstitial fluid. all these complex effects of 
histamine acting on different receptors where characterized by some guy 
named Lewis, who called it the triple response - starts with small local 
red spot at site of histamine injection (experimental, or via insect 
bite, or mast cell release) due to local dilation of blood vessels from 
H1 receptors/NO release. then you get large red flare, from continued 
vasodilation and poorly understood neural events, and probably it gets 
bigger and bigger due to H2 response. then you get a wheal - outer ring 
starts swelling - this is the vascular permeability, edema. 

histamine 
can also act on other smooth msucles, to produce contractions, but this 
is species specific and relates to H1/H2 distribution. H2 almost always 
associated with relaxation. spp with more H2 will have more relaxation.


histamine affects exocrine glands - causes marked increase in gastric 
acid secretion, often accompanied by increased secretion of pepsin.this 
isn't blocked by H1 antagonists. 

also stimulates salivary, pancreatic, 
intestinal, bronchial, and lacrimal secretions and maybe catecholamine 
release from adrenal medulla. pheochromocytoma patients get big BP rises 
when treated with histamine - seems paradoxical but mediated by 
catecholamine release.

we don't understand all of this but gastric acid 
secretion is well understood. histamine is involved in basal release and 
stimulated release. 

histamine is clearly involved in hypersensitivity 
and allergic responses, nasal and bronchial secretions are mediated by 
histamine. 

very abundant in brain - like serotonin, it's remarkable how 
much overlap there is b/w CNS effects - including control of sleep/wake 
cycles, depression, affective disorders, cerebral circulatory control, 
central control of water intake. probably interacts w/serotonin in 
important ways in brain. also involved in CNS control of reproduction , 
perhaps via controlling repro hormone release from pituitary.

drugs:

H1 
antagonists - seldane, hismanal - other antihistamines. very commonly 
used compounds for tx of colds, allergies,etc. usually well absorbed 
orally, metabolized by liver,excreted in urine. these were the first 
antihistamines made - described in 1937. main mechanism of action is on 
bronchial smooth muscle. they help prevent bronchoconstriction and reduce 
bronchial and nasal secretions - but they have no direct bronchodilator 
effect. they just prevent bronchoconstriction that would be induced by 
allergens or something. in this regard they are like bradykinin. but they 
are NOT bronchodilators like epinephrine. side effects - drowsiness, 
sedation. they get into CNS,produce sedation. not good. these properties 
probably result from their action in the brain. now we haev drugs that 
have poor entry into the CNS. and some of the meds used to treat colds at 
night take advantage of the sedative properties, and add some alcohol as 
well. these drugs also dry out mucous membranes. sometimes skin allergies 
develop following topical use.
another side effect - even H1 blockers 
have some GI effects - they can affect GI motility, and can even affect 
appetite. so sometimes antihistamines are prescribed to be taken only 
with food. 

H2 antagonists - cimetidine, etc.tagament, zantac, pepsin. 
structurally different from H1compounds - have big side chains on there. 
these are absorbed well orally, distribution pretty uniform but don't 
enter CNS due to big side chains. they are excreted in urine and feces. 
t1/2 20-100 minutes. these regulate acid secretion. toxicity - kidney is 
susceptible, perhaps due to renal histamine receptors. pulmonary - 
pleural effusion has occured with longterm use. these drugs are commonly 
used but not recommended for long term use. these drugs inhibit gastric 
acid secretion. normally pepsin falls in parallel, but sometimes thereare 
paradoxical increases in pepsin. also may decrease muscarinic receptor 
mediated acid secretion. basically, H2 receptor antagonist affects both 
basal and stimulated acid secretion directly, and affects gastrin 
mediated release, and perhaps vagally mediated. so it's good for use with 
gastric ulcers. many hypersecretory conditions like tumors of nonbeta 
pancreatic cells have been described, that cause increased gastrin 
release - H2 receptor antagonists reduce the resulting H+ secretion. a 
bunch of other conditions like basophilic leukemia, and hyperhistaminemia 
states - are also treated with these drugs.

H3 receptor - no drugs yet. 
should be eventually drugs that regulate histamine release.

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