Neuroactive effects of cotinine on the hippocampus: behavioral and biochemical parameters.

The present work evaluated the effects of nicotine (NIC), cotinine (COT), mecamylamine (MEC), methyllycaconitine (MLA) and dihydro-beta-eritroidine (DHßE) on memory extinction and the following biochemical parameters of the hippocampus: lipid peroxidation (LPO), antioxidant capacity (AC) and the phosphorylation of Extracellular-Signal-Regulated Kinase (ERK 1/2). Young male rats that were implanted bilaterally with cannulae were submitted to memory extinction tests sessions, and their hippocampi were dissected for biochemical assays. The extinction of fear memory was significantly improved by both nicotine and its metabolite. Cotinine significantly increased LPO, while nicotine significantly decreased it. Antioxidant capacity was increased by all treatments. Our results showed that cotinine, unlike nicotine, may increase oxidative stress in the hippocampus, but this increase depends upon the dose used and happens without causing corresponding impairments in cognitive function. Cotinine also increased the phosphorylation of ERK 1/2 in a similar fashion as nicotine. Considering these results, it is plausible to wonder to what extent nicotine-attributed effects are really due to the actions of this alkaloid and whether they could be due instead to cotinine or to cotinine-nicotine interactions within the brain.

Development of an anti-cotinine vaccine to potentiate nicotine-based smoking cessation strategies.

Nicotine replacement therapies (NRT) have limited success in smoking cessation. The efficacy of nicotine may be compromised by its main metabolite, cotinine. An anti-cotinine vaccine to remove this antagonism could enhance the efficacy of NRT. We show that cotinine is a weak nicotinic agonist and decreases responses to nicotine, consistent with antagonism through receptor desensitisation. trans-4-Thiol cotinine was coupled to tetanus toxoid, and rats immunised repeatedly. Vaccination raised antibodies specific for cotinine that do not recognise other metabolites or nicotine. Increased serum cotinine concentrations following nicotine administration indicate sequestration of cotinine by antibodies, encouraging further evaluation of this vaccine in behavioural models of nicotine addiction and relapse.

Evaluation of methoxsalen, tranylcypromine, and tryptamine as specific and selective CYP2A6 inhibitors in vitro.

CYP2A6 is the principle enzyme metabolizing nicotine to its inactive metabolite cotinine. In this study, the selective probe reactions for each major cytochrome P450 (P450) were used to evaluate the specificity and selectivity of the CYP2A6 inhibitors methoxsalen, tranylcypromine, and tryptamine in cDNA-expressing and human liver microsomes. Phenacetin O-deethylation (CYP1A2), coumarin 7-hydroxylation (CYP2A6), diclofenac 4'-hydroxylation (CYP2C9), omeprazole 5-hydroxylation (CYP2C19), dextromethorphan O-demethylation (CYP2D6), 7-ethoxy-4-trifluoromethylcoumarin deethylation (CYP2B6), p-nitrophenol hydroxylation (CYP2E1), and omeprazole sulfonation (CYP3A4) were used as index reactions. Apparent K(i) values for inhibition of P450s' (1A2, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1, and 3A4) activities showed that tranylcypromine, methoxsalen, and tryptamine have high specificity and relative selectivity for CYP2A6. In cDNA-expressing microsomes, tranylcypromine inhibited CYP2A6 (K(i) = 0.08 microM) with about 60- to 5000-fold greater potency relative to other P450s. Methoxsalen inhibited CYP2A6 (K(i) = 0.8 microM) with about 3.5- 94-fold greater potency than other P450s, except for CYP1A2 (K(i) = 0.2 microM). Tryptamine inhibited CYP2A6 (K(i) = 1.7 microM) with about 6.5- 213-fold greater potency relative to other P450s, except for CYP1A2 (K(i) = 1.7 microM). Similar results were also obtained with methoxsalen and tranylcypromine in human liver microsomes. R-(+)-Tranylcypromine, (+/-)-tranylcypromine, and S-(-)-tranylcypromine competitively inhibited CYP2A6-mediated metabolism of nicotine with apparent K(i) values of 0.05, 0.08, and 2.0 microM, respectively. Tranylcypromine [particularly R-(+) isomer], tryptamine, and methoxsalen are specific and relatively selective for CYP2A6 and may be useful in vivo to decrease smoking by inhibiting nicotine metabolism with a low risk of metabolic drug interactions.

Cotinine and nicotine inhibit each other's calcium responses in bovine chromaffin cells.

Cotinine is the major metabolite of nicotine. It has some biological activity, but its pathophysiological effects are largely unclear. We studied whether cotinine initiates calcium transients or affects those induced by nicotine. In bovine adrenal chromaffin cells labeled with the fluorescent calcium indicator Fura 2, cotinine (0. 32-3.2 mM) concentration-dependently increased the intracellular Ca(2+) concentration ([Ca(2+)](i)). The effect was abolished by omitting extracellular Ca(2+) during the stimulations. Also nicotinic receptor channel blockers hexamethonium (10 microM-1 mM) and chlorisondamine (100 microM), as well as a competitive nicotinic receptor antagonist dihydro-beta-erythroidine (10-100 microM), inhibited the response. Cotinine (0.32-3.2 mM) preincubation for 2 min inhibited both the nicotine-induced and the cotinine-induced increases in [Ca(2+)](i). Also nicotine (3.2-10 microM) inhibited the cotinine-induced increase in [Ca(2+)](i). Tetrodotoxin (1 microM) and thapsigargin (1 microM) pretreatments did not affect the responses to cotinine, while 300 nM nimodipine partially inhibited the cotinine-induced increase in [Ca(2+)](i). The results indicate that cotinine has nicotine-like effects on chromaffin cells. It may also desensitize the nicotinic cholinergic receptors, possibly by acting as a low-affinity agonist at these receptors.

(S)-(-)-Cotinine, the major brain metabolite of nicotine, stimulates nicotinic receptors to evoke [3H]dopamine release from rat striatal slices in a calcium-dependent manner.

Cotinine, a major peripheral metabolite of nicotine, has recently been shown to be the most abundant metabolite in rat brain after peripheral nicotine administration. However, little attention has been focused on the contribution of cotinine to the pharmacological effects of nicotine exposure in either animals or humans. The present study determined the concentration-response relationship for (S)-(-)-cotinine-evoked 3H overflow from superfused rat striatal slices preloaded with [3H]dopamine ([3H]DA) and whether this response was mediated by nicotinic receptor stimulation. (S)-(-)-Cotinine (1 microM to 3 mM) evoked 3H overflow from [3H]DA-preloaded rat striatal slices in a concentration-dependent manner with an EC50 value of 30 microM, indicating a lower potency than either (S)-(-)-nicotine or the active nicotine metabolite, (S)-(-)-nornicotine. As reported for (S)-(-)-nicotine and (S)-(-)-nornicotine, desensitization to the effect of (S)-(-)-cotinine was observed. The classic nicotinic receptor antagonists mecamylamine and dihydro-beta-erythroidine inhibited the response to (S)-(-)-cotinine (1-100 microM). Additionally, 3H overflow evoked by (S)-(-)-cotinine (10-1000 microM) was inhibited by superfusion with a low calcium buffer. Interestingly, over the same concentration range, (S)-(-)-cotinine did not inhibit [3H]DA uptake into striatal synaptosomes. These results demonstrate that (S)-(-)-cotinine, a constituent of tobacco products and the major metabolite of nicotine, stimulates nicotinic receptors to evoke the release of DA in a calcium-dependent manner from superfused rat striatal slices. Thus, (S)-(-)-cotinine likely contributes to the neuropharmacological effects of nicotine and tobacco use.