Table 1.

A summary of the literature on the heritable effects of paternal exposure to cigarette smoke or nicotine in animal models.

			Direct exposure of male founders			F1 (intergenerational effects)				F2 (transgenerational effects)
Exposure type	Length of exposure	Nicotine withdrawal prior to breeding	Behavior	Spermatozoa	Other effects	Behavior	Neurochemistry	Spermatozoa	Other effects	Behavior	Other effects
Cigarette Smoke: 2× daily for 1 h; Nicotine intraperitoneal, 0.05 mg/100 g 4×/day	5 weeks	No	Cigarette smoke increased depression like behavior and decreased locomotor activity. Nicotine exposure also increased depression like behavior but had no effect on locomotor activity. Neither exposure produced significant effect on anxiety, novel object recognition, or social behavior.	Enrichment of mRNA in the Wnt signaling pathway; increased WNT4 and DLV2 mRNA and protein expression in testis. Downregulation of Mmu-miRNA-15b with accompanying increase in methylation at CpG island shore regions.	Effects of nicotine exposure, but not cigarette smoke exposure, were examined on WNT4 and DLV2 expression in brain to compare with expression in testes. Expression was not significantly changes in the brain	Exposures to cigarette smoke and nicotine both produced significant decrease in depression like behavior, and increases in social behavior and locomotor activity.	N/A	No significant effect on Mmu-miRNA-15b expression	Nicotine exposure increased WNT4 and DVL2 protein expression in the brain. WNT4 mRNA but not DVL2 mRNA expression was increased significantly as well. Expression of Mmu-miRNA-15b, which represses WNT4 expression, was downregulated significantly in the brain. The CpG island shore regions of this miRNA showed significant hypermethylation.	No significant effect on depression like behavior, anxiety, novel object recognition, locomotor activity, and social behavior.	No significant effect on WNT and DLV4 expression
Nicotine 200 μg/ml in drinking water sweetened with saccharin	5 weeks	Yes	No significant effects on anxiety-like behavior	N/A	N/A	No significant effect on nicotine-induced suppression of locomotor activity in novel environment or on nicotine self-administration. However, paternal nicotine exposure “protected” the mice from lethal effects of high-dose nicotine, if they had been exposed previously to chronic sub-lethal doses, but not upon the first administration of the lethal dose.	N/A	N/A	Male but not female mice showed increased resistance to nicotine and cocaine toxicity and upregulation of xenobiotic metabolism. Hepatocytes showed reduced apoptosis and necrosis following cocaine exposure.	N/A	N/A
Nicotine 200 μg/ml in drinking water	12 weeks	No	No significant effect on locomotor activity, spatial working memory or object based attention	Increased global DNA methylation. Hypomethylation of DNA at dopamine D2 receptor promoter region. No significant change in DNA in methylation of dopamine D1, D3, D4, or D5 receptor promoter regions	N/A	Both male and female mice showed significant hyperactivity and reversal learning deficit. Male mice also showed significant attention deficit. No significant effects on working memory in male or female mice.	Male mice showed significant decreases in striatal tissue content of dopamine and its metabolites as well as significant increase in frontal cortical tissue content of norepinephrine. Male mice also showed significant decreases in mRNA expression for dopamine D1, D2, and D4 receptors in the striatum without significant changes in dopamine D3 and D5 receptor mRNA. Female mice showed significant increase in striatal dopamine D5 receptor mRNA without significant changes in dopamine D1, D2, D3, or D4 receptor mRNA. There was no change in striatal tissue content of dopamine or dopamine metabolites nor any significant change in frontal cortical tissue content of norepinephrine.	N/A	N/A	Male F2 mice from female F1 founders showed significant reversal learning deficit but no significant change in locomotor activity, working memory or attention. Male F2 mice from F1 male founders and female F2 mice from male or female F1 founders did not significant changes in locomotor activity, working memory, or attention.	N/A
Nicotine; 12.6 mg/day/x4 via subcutaneous mini pump	28 days	Yes	N/A	N/A	N/A	Both male and female mice showed significant decreases in nicotine self-administration and significant increases in contextual and cued fear responses.	Both male and female mice showed significant upregulation of hippocampal nicotinic acetylcholine receptor ligand binding and significant reduction in evoked hippocampal cholinergic currents.	N/A	Both male and female mice showed significant changes in DNA methylation and mRNA expression of genes related to neural development and plasticity in the hippocampus.	Contextual and cued fear responses were significantly upregulated	N/A
Reference	Exposure type	Length of exposure	Nicotine withdrawal prior to breeding	Behavior	Spermatozoa	Other effects	Behavior	Neurochemistry	Spermatozoa	Other effects	Behavior	Other effects
Zhang et al. (2020)	Nicotine 0.05 mg/100 g x4/day intraperitoneal	5 weeks	No	N/A	Total DNA methylation rate of dopamine transporter gene was upregulated	N/A	Male mice were significantly hyperactive	Hippocampal dopamine release and tissue content were significantly upregulated in male and female mice along with significant downregulation of dopamine transporter mRNA and protein expression. Total DNA methylation rate of dopamine transporter gene was upregulated along with activation of the dopamine D2 receptor/AKT/GSKα/β pathway.	N/A		N/A	N/A
Murphy et al. (2020)	Cigarette smoke exposure 2.5 h/day for 5 days a week	60 days	Yes	N/A	Significant up- and downregulation of DNA methylation occurred depending on the methylation status of the DNA at baseline. The baseline methylation status also impacted the rate at which the cigarette-smoke-induced changes were lost following cessation of the exposure. DNA methylation at regions that showed extreme hyper- and hypo-methylation at baseline was less likely to be affected by the smoke exposure and when changes did occur these regions were more likely to recover upon withdrawal of exposure.	N/A	N/A	N/A	There was no significant correlation between DNA methylation patterns in the spermatozoa of directly cigarette smoke exposed mice (founders) and spermatozoa of the F1 mice.	Significant changes in DNA methylation in over 28,000 regions in the prefrontal cortex in male mice and differential expression of 134 genes associated with oxidative stress.	N/A	N/A
Zhang et al. (2021)	Nicotine (0.01% (v/v) in saline (5 ml/kg) intraperitoneal; 4x/day	5 weeks	No	N/A	Downregulation of mmu-miRNA-15b via hypermethylation of CpG island shore regions.	N/A	N/A	N/A	N/A	Upregulation of mRNA and protein levels of α-SMA and Col1α1 in F1 male offspring The increased hepatic fibrosis was a result of upregulation of Wnt4, Dvl2 and β-catenin and downregulation of Gsk-3β. Downregulation of mmu-miRNA-15b via hypermethylation of DNA in F1 liver (i.e. activation of Wnt pathway).	N/A	No change in Wnt4 mRNA and protein expression, mmu-miR-15b expression, or methylation state of mmu-miR-15b in liver.
Zeid et al. (2021)	Nicotine; 12.6 mg/day/x4 via subcutaneous mini pump	28 days	Yes	N/A	N/A	N/A	No change in sensitivity to nicotine-induced hypo locomotion in F1 male and female offspring.	N/A	N/A	Reduced sensitivity to nicotine-induced hypothermia, and delayed nicotine clearance in F1 male mice. Reduced basal corticosterone in F1 female offspring.	N/A	N/A

Data and conclusions from each publication are presented separately for the exposed founder, his offspring (F1) and the second generation (F2). The findings on behavior as well as neurochemistry, gene expression, and epigenetic changes in the brain and spermatozoa are presented