MODES OF NEUROPHARMACOLOGICAL ACTIONS OF TAURINE AND ITS POTENTIAL THERAPEUTIC USAGE Shailesh P. Banerjee, Christopher Y. Chan & Eitan Friedman CUNY School of Medicine, New York, NY 10031

THIS PRESENTATION WILL BE A REVIEW OF OUR RESEARCH ON TAURINE HOW AND WHY WE GOT INVOLVED IN STUDIES RELATED TO TAURINE? ANTIPSYCHOTIC-INDUCED CATALEPSY a. Haldol (haloperidol)-induced catalepsy b. Protection by taurine COCAINE-INDUCED NEUROTOXICITY a. Effects of perinatal cocaine exposure b. Endogenous taurine’s antagonsim of cocaine-induced neurotoxicity TAURINE’S ANTI-ADDICTING ACTIVITY a. Inhibition of sensitization of locomotor activity induced by cocaine b. Inhibition of cocaine-induced conditioned place preferance TAURINE’S INTERACTION WITH THE GLUTAMATE NMDA RECEPTOR a. Electrophysiological analysis b. Receptor binding assays c. Analysis of NMDA receptor subunits by immuno-blotting 6. MECHANISMS FOR TAURINE’S MODES OF ACTIONS a. Glycine-gated Chloride channel b. Taurine-gated Chloride channel c. Intracellular Calcium d. NMDA Glutamate receptor inhibition

1. Our Interest in Taurine

2. ANTIPSYCHOTIC-INDUCED CATALEPSY i 2. ANTIPSYCHOTIC-INDUCED CATALEPSY i. Metoclopramide sensitizes haloperidol-induced catalepsy Solvent Metoclopramide (5 mg/kg) Metoclopramide (10 mg/kg) Haloperidol (0.5 mg/kg) n=8 Twenty-four rats were divided into three groups with eight animals in each group. The first group received solvent and served as control. Groups 2 and 3 received 5 or 10 mg/kg of metoclopramide, respectively. A week later all 24 rats received 0.5 mg/kg haloperidol (I). Administration of 0.5 mg/kg haloperidol was repeated in weeks two (II) and three (III). The data are expressed as mean ± S.E.M. Statistical differences were evaluated using One-way ANOVA followed by Tukey test for multiple comparisons. *, P < 0.05; **, P< 0.01 when compared with respective value in comparison group. ------------------------------------------------------------------------------------------ Results Seven days after a single dose of 0.5 mg/kg haloperidol animals showed moderate but significant degree of catalepsy compared with control animals that received an equal volume of solvent. The intensity of catalepsy was significantly enhanced at weeks 2 and 3 following weekly administration of the same dose of haloperidol. These results are similar to our previously reported observation on cataleptic effects of haloperidol (Lidsky et al. 1994, 1995). In sharp contrast, a week after a single injection of 5 mg/kg or 10 mg/kg of metoclopramide no catalepsy was observed. Interestingly, when rats pretreated with 5 or 10 mg/kg of metoclopramide were given 0.5 mg/kg of haloperidol seven days later, animals exhibited significantly higher degree of catalepsy compared to solvent plus haloperidol treated rats. Further, weekly administration of the same dose of haloperidol showed significant enhancement of catalepsy in the metoclopramide-pretreated groups compared to solvent plus haloperidol group at weeks two and three. The degree of catalepsy was significantly greater in animals receiving 10 mg/kg metoclopramide compared to those receiving 5 mg/kg metoclopramide at each of the three experimental time points. Thus, metoclopramide was found to be effective in sensitizing haloperidol-induced catalepsy. From Agovic et al., 2008. E. J. Pharm.

ANTIPSYCHOTIC-INDUCED CATALEPSY ii. NMDA and D2 receptor binding [3H-Spiroperidol binding] [3H-MK 801 binding] Agovic et al., 2008. E. J. Pharm.

ANTIPSYCHOTIC-INDUCED CATALEPSY HYPOTHESIS Antipsychotic-induced catalepsy involves Augmentation of NMDA-mediated glutamatergic transmission; Inhibition of dopamine D2 receptor mediated- transmission; and Unchanged dopamine D1 receptor-mediated transmission.

TWO POSSIBLE MECHANISMS FOR THE DEVELOPMENT OF CATALEPSY 1. Impairment Of Neurotransmitter Functions 2. Neurodegeneration

Taurine’s protection against haldol-induced catalepsy Haldol only Haldol + taurine Lidsky et al., 1995 Brain Research

Taurine’s protection against haldol-induced neurodegeneration b From Lidsky et al., 1995 Brain Research

Summary Antipsychotic-induced catalepsy involves 1. Impairment Of Neurotransmitter Functions & 2. Neurodegeneration TAURINE PREVENTS BOTH OF THESE EFFECTS.

3. COCAINE-INDUCED NEUROTOXICITY Effects of perinatal cocaine exposure From Yablonski-Alter et al., 2005 JPET

COCAINE-INDUCED NEUROTOXICITY B. Endogenous taurine’s antagonism of cocaine-induced neurotoxicity - I

COCAINE-INDUCED NEUROTOXICITY B. Endogenous taurine’s antagonism of cocaine-induced neurotoxicity - II

MECHANISMS FOR PROTECTION AGAINST COCAINE-INDUCED TOXICITY Inhibition of glycine release Stimulation of taurine release

4. TAURINE’S ANTI-ADDICTING ACTIVITY A. INHIBITION OF SENSITIZATION OF LOCOMOTOR ACTIVITY INDUCED BY COCAINE

TAURINE’S ANTI-ADDICTING ACTIVITY B. INHIBITION OF COCAINE-INDUCED CONDITIONED PLACE PREFERANCE

Taurine (2mM) inhibited the late part of N2. Portion removed by taurine ..but N1 is spared 17

Does co-application of the slice with the NMDA antagonist, APV, prevent the inhibition caused by taurine. APV = 2 amino, 5-phosphonovalerate

b c a Taurine effect was prevented in slices treated with APV APV+Tau 50 100 % inhibition of Control amplitude 73.28% 73.15% (1) Con (2) APV (3) APV+Tau N2 a N = 5 Taurine effect was prevented in slices treated with 100 mM DL-AVP, a specific NMDA receptor antagonist. 19

[3H] MK801, [3H] spermidine, [3H] taurine binding in rat cortical membranes specific ligand membrane binding was performed in the presence of 30mM Glycine

Spermine dose-dependently enhanced [3H] MK801 binding

Taurine dose-dependently reduced spermine- enhanced [3H]MK801 binding 30mM glycine + 100mM spermine. Note that taurine had no direct effect on [3H] MK801 binding per se (not illustrated).

Taurine reduced the affinity of glycine for the NMDA receptor by about 10 fold

Polyamines displace both 3[H]spermine and 3[H]taurine from cortical tissue

2mM Tauine co-applied with 10 mM Ro25-6981 induced a slight additional inhibition, much less than the 33 +/- 4% inhibition caused by taurine alone. 2 mM taurine +10mM R025-6981 [Ro25-6981], mM % inhibition of the N2 component P < 0.1 2 mM taurine alone

The inhibition of the N2 response component by 2mM taurine is progressively replaced by increasing doses of Ro25-6981: No additive interaction between the 2 modulators P < 0.01 Tau P < 0.05 % inhibition of the N2 response component [Ro25-6981], mM, in the absence or presence of 2mM taurine

A hypothetical additive interaction suggests independent mechanisms by the 2 modulators would produce an equal upward shift of the % inhibition regardless of Ro25-6981 concentrations. Predicted action of Ro25 + Taurine [Ro25-6981] Action of Ro25 % inhibition of N2 component

Long term taurine treatment increases the membrane expression of NMDA receptor subunit GluN2B Optical Intensity (Arbitrary Units) Long term (30d) taurine treatment increases the expression of NMDA receptor subunit GluN2B: significant increase in membrane expression of GluN2B subunit (p<0.05, unpaired T-test).

Long term taurine treatment increases the membrane expression of NMDA receptor subunit GluN1

Long term taurine treatment does not alter the membrane expression of AMPA receptor subunit GluR1 Optical Intensity (Arbitrary Units) Long term (30d) taurine treatment does not alter the synaptic membrane expression of AMPA receptor subunit GluR1:

Long term taurine treatment reduces the membrane expression of AMPA receptor subunit GluR2 Optical Intensity (Arbitrary Units) Long term (30d) taurine treatment reduces the synaptic membrane expression of AMPA receptor subunit. p<0.05, unpaired T-test, N=5.

5. MECHANISMS FOR TAURINE’S MODES OF ACTIONS Glycine-gated chloride channel in the spinal cord (Borghese et al., 2012 JPET). Taurine-activated chloride influx (Yarbrough et al., 1981 JPET). Decreasing the intracellular level of free calcium (El Idrissi and Trenkner, 2003 Adv.Exp.Med.Biol.). Inhibition of NMDA glutamate receptor subtype (Chan et al., 2012 These Proceedings).

Taurine’s Multiple Activities

Acknowledgements: Ted I. Lidsky, Ph.D. Elena Yablonski-Alter, Ph.D. Mervan Agovic, Ph.D. Kalindi Bakshi, Ph.D. Eleonora Gashi, D.O. Brice le Francois, Ph.D. Andre Ragnauth, Ph.D. Louis Vidal, Ph.D. Eitan Friedman, Ph.D.

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Extra slides

TAURINE’S ANTI-ADDICTING ACTIVITY C. CHRONIC TAURINE INHIBITS COCAINE-INDUCED DOPAMINE RELEASE A.T. (After taurine)

Polyamines and Taurine compete for a common binding site 3H-Taurine competition 3H-Spermidine competition Crude membrane preparations were incubated with tritiated taurine (1μM) or spermidine (0.25 μM) in the presence of 30 µM glycine and non radioactive spermidine, spermine or taurine.