1. Calcium Channel Blockers
October 3, 2007
Frank F. Vincenzi

2. Ca signalling and Ca channel blockers: The fundamental information
extracellular
intracellular = 2+ -7 2+ -3 Ca
10 M Ca =
10 M

3. Calcium
The FIRST second messenger fundamental intracellular messenger for a wide variety of physiological responses in essentially all cell types (muscle tension, neurotransmitter/hormone release, production of inflammatory mediators)
The LAST second messenger accumulation of intracellular Ca2+ is a final common pathway in both apoptotic and necrotic cell death

4. Ca signalling: Two fundamental mechanisms
Influx of Ca across the plasma membrane
voltage-operated Ca channels (VOCs)
receptor-operated Ca channels (ROCs)
Internal release of Ca from intracellular stores (sarcoplasmic/endoplasmic reticulum) (SER)
Ca-induced Ca release (‘trigger Ca’)
Inositol tris-phosphate( IP3) induced Ca release
Combinations of the above mechanisms

5. Intracellular Calcium Release
Inositol tris-phosphate (IP3)
Present in almost all cells. IP3 acts on sarcoplasmic/endoplasmic reticulum receptors (IP3Rs)
IP3 may sensitize Ca-induced Ca release
Calcium-induced calcium release
Mainly in striated muscle
Ryanodine receptors (RyRs)
RyR1 (predominantly in skeletal muscle)
[Mutation associated with malignant hyperthermia]
RyR2 (predominantly in cardiac muscle)
RyR3 (predominantly in brain & non-muscle)

6. Agonist Receptor operated Ca channels mediate some agonist actions Ca+ Ca 2+ Ca 2+ Ca 2+ Na
Agonist 2+ Ca 2+ Ca R R
sarcolemma
ATP O O C C Ca 2+ 2+ Ca 2+ Ca Ca 2+
RyR
Receptor operated
Ca channels mediate
some agonist actions Ca 2+ Ca 2+ Ca 2+ SR

7. Ca signalling: Three examples
Histamine on H-1 receptor activates phospholipase C and releases inositol trisphosphate (IP3), IP3 acts on SR/ER IP3 receptors causing Ca release (CHEMICAL)
Norepinephrine on beta-1 receptors in heart couple to G protein and activate adenylyl cyclase; increase cAMP, activate protein kinase A (PKA), phophorylate a cytoplasmic domain of the Ca channel, increase Ca current; PKA also phosphorylates phospholamban, an SR protein and increases the activity of the SERCA pump; increase Ca storage and release (CHEMICAL/ELECTRICAL)
Cell depolarization activates voltage operated Ca channels (VOCs) in the plasma membrane, promoting influx of Ca into the cell cytoplasm, may be amplified by Ca induced Ca release (ELECTRICAL)

8. histamine Ca Ca Ca Na Ca sarcolemma Ca Ca Ca Ca RyR Ca Ca Ca SR PLC+ Ca 2+ Ca 2+ Ca 2+ Na
histamine Ca 2+
sarcolemma
ATP
PLC
DAG Ca 2+
IP3 Ca 2+ Ca 2+ Ca 2+
RyR IP R 3 Ca 2+
Phospholipase C and
inositol tris phosphate
mediate some agonist
actions by promoting
intracellular Ca release Ca 2+ Ca 2+ SR

9. AC Ca Ca Ca Na Ca sarcolemma Ca Ca Ca Ca Ca RyR Ca Ca Ca Ca Ca Ca Ca+ Ca 2+ Ca 2+ Ca 2+ Na Ca 2+ V O C V O C
sarcolemma
Beta-1 Receptor
ATP AC G Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+
RyR Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+

10. norepinephrine AC cAMP PKA cAMP and protein kinase A mediate some+ Ca 2+
norepinephrine Ca 2+ Ca 2+ Na Ca 2+ V O C V O C
sarcolemma
Beta-1 Receptor
ATP AC G P Ca 2+ Ca 2+ Ca 2+
cAMP Ca 2+ Ca 2+
RyR
PKA Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+
cAMP and protein
kinase A mediate some
of the positive inotropic
effects of catecholamines
by increasing Ca influx
and intracellular Ca storage
and release Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ P

11. Ca regulation of muscle tension: different in different types of muscle
Reversible ionic binding of Ca to the Ca binding protein, troponin C (Ca/TNC disinhibits actin/myosin ATPase)
skeletal myo (force varied by recruitment)
cardiac myo (force varied by cellular regulation)
Covalent modification of myosin light chain (Ca binds to the Ca binding protein, calmodulin CaM, CaM binds to MLCK, MLCK phosphorylates MLC, activates ATPase - tension)
smooth myo (force varied by level of MLC phosphorylation)

12. The calcium cycle in HEART muscle+ Ca 2+ Ca 2+ Ca 2+ Na 2+ 2+ Ca 2+ Ca Ca V O C V O C
sarcolemma
ATP
Ca channel, DHPR Ca 2+ 2+ 2+ Ca Ca Ca 2+
RyR Ca 2+ Ca 2+ Ca 2+ SR

13. The calcium cycle in SMOOTH muscle

14. The calcium cycle in SKELETAL muscle2+ Ca 2+ 2+ 2+ Ca Ca 2+ Ca
sarcolemma
Ca “channel”, DHPR 2+ Ca Ca 2+
T-tubule
RyR Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ Ca 2+ SR

15. Ca regulation in muscle: variations on a few themes
VOC
ROC
ROC
PMCA
Na/Ca 2+ SR Ca
exchange + Na
actin
Na/K
myosin
mitochondria
pump + K

16. Muscle control: differential sensitivity to Ca channel blockers (CEBs)

17. Overview of Plasma Membrane Ca Channels
Voltage operated channels (VOCs) L-type, N-type, T-type….etc.
Receptor operated channels (ROCs) coupled to diverse receptors in the same cell
Mechanically operated channels (MOCs) not widely appreciated (yet)

18. Voltage operated Ca Channels
L-type (‘long’, important in heart & smooth muscle)
N-type (‘nerve’, important in conduction & synaptic transmission)
T-type (‘transient’, heart & smooth muscle)

19. Calcium Channel Blockers
Currently marketed Ca channel blockers all block L-type channels.
Other terminology for these drugs: Calcium entry blockers (CEBs) Slow channel blockers “Calcium antagonists” (ugh!)
(why NOT call them calcium antagonists?)

20. Chemical classifications of Ca channel blockers
dihydropyridines e.g., nifedipine (Procardia®) (have given the name to voltage operated Ca signaling proteins - ‘the dihydropyridine receptor’
benzothiazepines e.g., diltiazem (Cardizem®)
phenylalkylamines e.g., verapamil (Isoptin®, Calan®)

21. In General Calcium Channel Blockers Are:
Lipid soluble
Rapidly absorbed
Largely protein bound
Of low bioavailability
Metabolized by the liver

22. Calcium Channel Blockers: Site and Mechanism of Action
L-type voltage operated channels (VOCs) and receptor operated channels (ROCs)
Binding to Ca channel protein (slightly different site for each class):
inhibits ionophoric activity of affected channels (all agents)
delays recovery of the affected voltage operated channels (verapamil > diltiazem >> nifedipine)

23. Calcium Channel Blockers: Frequency Dependence
Verapamil (and to some extent, diltiazem) selectively inhibits rapid cardiac rhythms by delaying recovery of calcium channels
Dihydropyridines, such as nifedipine, have little or no effect on cardiac Ca channels in vivo

24. Calcium channel blockers: potencies for different tissues compared
Smooth muscle dihydropyridines (e.g., nifedipine) >> verapamil ~ diltiazem
Heart
SA & AV nodes verapamil ~ diltiazem >> nifedipine
myocardium verapamil > diltiazem >> nifedipine

25. Clinical Effects of Ca Channel Blockers
Relax arteriolar smooth muscle, little or no venodilation
Verapamil & diltiazem also depress SA and AV nodes and depress myocardial contractility to some extent.
Dihydropyridines do not affect heart in vivo in normal doses
Note: little or no effect on nervous system or skeletal muscle (no block of N-type Ca channels, and internal cycling of Ca, respectively)

26. Calcium Channel Blockers: Indications
Angina (multiple mechanisms)
Atrial arrhythmias (AV node conduction depends on ‘Ca action potentials’ - some Ca channel blockers increase AV node refractory period)
Hypertension (decrease vascular resistance) ( ± nifedipine for malignant hypertension) (sodium nitroprusside is a drug of choice here) (Raynaud’s syndrome?, asthma?, atherosclerosis?, reperfusion injury?)

27. nifedipine: indications
Angina, variant angina
Hypertension
Off label aortic regurgitation, diabetic nephropathy, hiccups, hypertensive emergency/urgency, prophylaxis of migraine, premature labor (FDA-use-in-pregnancy C)

28. nifedipine: selected adverse reactions
Hypotension
Peripheral edema
Flushing, headache

29. diltiazem: indications
Angina, variant angina
Atrial flutter, atrial fibrillation
Paroxysmal supraventricular tachycardia
Hypertension - slow release formulations only
Off label Cardiomyopathy, diabetic nephropathy, proteinuria, prophylaxis of migraine

30. diltiazem: selected adverse reactions
Depression of cardiac muscle
AV nodal blockade
Hypotension
Peripheral edema
Flushing, headache

31. verapamil: indications
Angina, unstable angina, variant angina
Atrial flutter, atrial fibrillation
Paroxysmal atrial tachycardia (prevent and Tx)
Hypertension
Off label claudication, prophylaxis of migraine, mania

32. verapamil: selected adverse reactions
Depression of cardiac muscle
AV nodal blockade
Hypotension
Peripheral edema
Flushing, headache
Constipation

33. Psaty et al., (1995)* Use of short acting (emphasis added) Ca channel blockers, especially in high doses, and particularly when combined with diuretics, was associated with an increased risk of myocardial infarction. (None of the therapies monitored decreased MI risk significantly) Cochrane, different result with long acting CEBs
*JAMA 274: , 1995

34. Scarlett O’Brian
Scarlett O’Brian is a 38 year old female who was diagnosed with mild hypertension. She was given a prescription for verapamil (80 mg, TID, #100).
Two days later she called her boyfriend (a prominent [married] business executive) to an urgent meeting to ‘sort out their relationship’.
The boyfriend later testified that they had a ‘couple of drinks’ while he explained that he was not going to leave his wife.

35. Scarlett O’Brian collapses
As they continued to talk, Scarlett began to feel poorly. She fainted, but appeared to be breathing adequately. Over the course of the next hour, while lying down, she became worse with a pounding headache, profound ‘weakness’ and inability to stand.
The boyfriend delayed calling 911 for fear of making their relationship public. When she became unconscious, he finally called. By the time EMTs arrived she was in complete heart block. Efforts to restore an effective cardiac rhythm failed.

36. Scarlett O’Brian, what happened?
Did she die of some underlying disease?
Did she die of alcohol poisoning?
Did she die because of verapamil?

37. Scarlett O’Brian, Autopsy
Body is that of a well developed, well nourished female (60 kg).
No structural cause of death could be identified.
Gastric contents contained partially disintegrated tablets (yellow)
Toxicology (blood) identified: caffeine - trace ethanol mg/l (0.035 BAC) verapamil - 23 mg/L

38. If taken as prescribed, what was the predicted average steady state concentration of verapamil in Scarlett?
CL verapamil = 15 mL/kg/min = 54 L/h in 60 kg pt.
Rx: 80 mg TID = 240 mg/24 h = 10 mg/h
Css = (10 mg/h) * (0.22) / (54 L/h) = 0.04 mg/L (equivalent to 40 ng/mL)

39. Verapamil, some useful numbers
NORMALLY, a large first-pass effect, 22% bioavailability (metabolized in the liver to norverapamil)
Urinary excretion < 3%
Plasma binding = 90%
CL = ‘15 mL/min/kg’ (15 mL*min-1*kg-1)
Vd = 5 L/kg
Half life = 4 hours
Effective concs. = 120 ng/mL
Toxic concs. ?

40. Scarlett, more questions
Was Scarlett a poor metabolizer?
Suppose she had metabolized none of the verapamil, how much should be in her body? 3 days * 240 mg/day = 720 mg
How much verapamil was in the prescription?
100 tablets * 80 mg/tab = 8000 mg
Even if Scarlett had taken all 100 tablets at once, if the drug was absorbed and metabolized normally only about 1600 mg should have been present.

41. Scarlett, the numbers tell the story
How much verapamil should she have taken? (three days time 240 mg/day = 720 mg)
Theoretical Vd of verapamil in Scarlett’s body? (60 kg * 5 L/kg = 300 L total Vd)*
Blood conc at autopsy = 23 mg/L
IF distributed in total Vd, then the total verapamil in Scarlett’s body:
300 L * 23 mg/L = 6900 mg.

42. Scarlett O’Brian: A suicide gesture gone bad
Scarlett probably took most or all of the prescription of verapamil shortly before her boyfriend arrived.
The drug was rapidly absorbed and saturated first pass mechanisms (bioavailability approached 1).
Hypotension and heart block were produced as the concentration of verapamil increased.
The delay in calling 911 probably turned a suicide gesture into a suicide.