Epigenetic factors affecting female rat sexual behavior

This 2018 Baltimore/Montreal rodent study found:

“If sexually naïve females have their formative sexually rewarding experiences paired with the same male, they will recognize that male and display mate-guarding behavior towards him in the presence of a female competitor. Female rats that display mate-guarding behavior also show enhanced activation of oxytocin and vasopressin neurons in the supraoptic and paraventricular hypothalamic nucleus.

We examined the effect of a lysine-specific demethylase-1 inhibitor to block the action of demethylase enzymes and maintain the methylation state of corresponding genes. Female rats treated with the demethylase inhibitor failed to show any measure of mate guarding, whereas females treated with vehicle displayed mate guarding behavior. Demethylase inhibitor treatment also blocked the ability of familiar male cues to activate oxytocin and vasopressin neurons, whereas vehicle-treated females showed this enhanced activation.”

General principles and their study-specific illustrations were:

Histone modifications are a key element in gene regulation through chromatin remodeling. Histone methylation / demethylation does not have straightforward transcriptional outcomes as do other histone modifications, like acetylation, which is almost invariably associated with transcriptional activation.

What is of vital importance in regards to histone methylation / demethylation is the pattern of methylation that is established. Patterns of methylation incorporate both methylated and demethylated residues, and are what ultimately play a role in transcriptional outcomes.

In the present study, inhibiting LSD1 demethylase enzymes disrupted the ability of cells to properly establish histone methylation / demethylation patterns, thus creating a deficit in the cells’ ability to transcribe the gene products necessary for the enhanced induction of OT, AVP, and the subsequent mate-guarding behaviors we observed. This study is the first to demonstrate a definitive role of epigenetic histone modifications in a conditioned sexual response.”

https://www.sciencedirect.com/science/article/pii/S0031938418303421 “Inhibition of lysine-specific demethylase enzyme disrupts sexually conditioned mate guarding in the female rat” (not freely available)


Hijacking the epigenetic clock paradigm

This 2018 German human study’s last sentence was:

“Additionally we found an association between DNAm [DNA methylation] age acceleration and rLTL [relative leukocyte telomere length], suggesting that this epigenetic clock, at least partially and possibly better than other epigenetic clocks, reflects biological age.”

Statements in the study that contradicted, qualified, and limited the concluding sentence included:

“The epigenetic clock seems to be mostly independent from the mitotic clock as measured by the rLTL.

It could be possible that associations are confounded due to short age ranges or non-continuous age distribution, as displayed in the BASE-II cohort (no participants between the age of 38 and 59 years). [see the below graphic]

The BASE-II is a convenience sample and participants have been shown to be positively selected with respect to education, health and cognition.

Samples in which DNAm age and chronological age differed more than three standard deviations from the mean were excluded (N=19).

While the original publication employed eight CpG sites for DNAm age estimation, we found that one of these sites did not significantly improve chronological age prediction in BASE-II. Thus, we reduced the number of sites considered to seven in the present study and adapted the algorithm to calculate DNAm age.

  • Horvath described a subset of 353 methylation sites predicting an individual’s chronological age with high accuracy..
  • Even though the available methods using more CpG sites to estimate DNAm age predict chronological age with higher accuracy..
  • It is not clear how much of the deviation between chronological age and DNAm age reflects measurement error/low number of methylation sites and which proportion can be attributed to biological age.

Due to the statistical method employed, we encountered a systematic deviation of DNAm age in our dataset.”

Findings that aren’t warranted by the data is an all-too-common problem with published research. This study illustrated how researcher hypothesis-seeking behavior – that disregarded what they knew or should have known – can combine with a statistics package to produce almost any finding.

It reminded me of A skin study that could have benefited from preregistration that made a similar methodological blunder:

The barbell shape of the subjects’ age distribution wouldn’t make sense if the researchers knew they were going to later use the epigenetic clock method.

The researchers did so, although the method’s instructive study noted:

“The standard deviation of age has a strong relationship with age correlation”

and provided further details in “The age correlation in a data set is determined by the standard deviation of age” section.

Didn’t the researchers, their organizations, and their sponsors realize that this study’s problematic design and performance could misdirect readers away from the valid epigenetic clock evidence they referenced? What purposes did it serve for them to publish this study?

https://academic.oup.com/biomedgerontology/advance-article-abstract/doi/10.1093/gerona/gly184/5076188 “Epigenetic clock and relative telomere length represent largely different aspects of aging in the Berlin Aging Study II (BASE-II)” (not freely available)

The role of recall neurons in traumatic memories

This 2018 Swiss rodent study found:

“Our data show that:

  • A subset of memory recall–induced neurons in the DG [dentate gyrus] becomes reactivated after memory attenuation,
  • The degree of fear reduction positively correlates with this reactivation, and
  • The continued activity of memory recall–induced neurons is critical for remote fear memory attenuation.

Although other brain areas such as the prefrontal cortex and the amygdala are likely to be implicated in remote fear memories and remain to be investigated, these results suggest that fear attenuation at least partially occurs in memory recall–induced ensembles through updating or unlearning of the original memory trace of fear.

These data thereby provide the first evidence at an engram-specific level that fear attenuation may not be driven only by extinction learning, that is, by an inhibitory memory trace different from the original fear trace.

Rather, our findings indicate that during remote fear memory attenuation both mechanisms likely coexist, albeit with the importance of the continued activity of memory recall–induced neurons experimentally documented herein. Such activity may not only represent the capacity for a valence change in DG engram cells but also be a prerequisite for memory reconsolidation, namely, an opportunity for learning inside the original memory trace.

As such, this activity likely constitutes a physiological correlate sine qua non for effective exposure therapies against traumatic memories in humans: the engagement, rather than the suppression, of the original trauma.”

The researchers also provided examples of human trauma:

“We dedicate this work to O.K.’s father, Mohamed Salah El-Dien, and J.G.’s mother, Wilma, who both sadly passed away during its completion.”

So, how can this study help humans? The study had disclosed and undisclosed limitations:

1. Humans aren’t lab rats. We can ourselves individually change our responses to experiential causes of ongoing adverse effects. Standard methodologies can only apply external treatments.

2. It’s a bridge too far to go from neural activity in transgenic mice to expressing unfounded opinions on:

“A physiological correlate sine qua non for effective exposure therapies against traumatic memories in humans.”

Human exposure therapies have many drawbacks, in addition to being applied externally to the patient on someone else’s schedule. A few others were discussed in The role of DNMT3a in fear memories:

  • “Inability to generalize its efficacy over time,
  • Potential return of adverse memory in the new/novel contexts,
  • Context-dependent nature of extinction which is widely viewed as the biological basis of exposure therapy.”

3. Rodent neural activity also doesn’t elevate recall to become an important goal of effective human therapies. Clearly, what the rodents experienced should be translated into human reliving/re-experiencing, not recall. Terminology used in animal studies preferentially has the same meaning with humans, since the purpose of animal studies is to help humans.

4. The researchers acknowledged that:

“Other brain areas such as the prefrontal cortex and the amygdala are likely to be implicated in remote fear memories and remain to be investigated.”

A study that provided evidence for basic principles of Primal Therapy determined another brain area:

“The findings imply that in response to traumatic stress, some individuals, instead of activating the glutamate system to store memories, activate the extra-synaptic GABA system and form inaccessible traumatic memories.”

The study I curated yesterday, Organ epigenetic memory, demonstrated organ memory storage. It’s hard to completely rule out that other body areas may also store traumatic memories.

The wide range of epigenetic memory storage vehicles is one reason why effective human therapies need to address the whole person, the whole body, and each individual’s entire history.

http://science.sciencemag.org/content/360/6394/1239 “Reactivation of recall-induced neurons contributes to remote fear memory attenuation” (not freely available)

Here’s one of the researchers’ outline:

This post has somehow become a target for spammers, and I’ve disabled comments. Readers can comment on other posts and indicate that they want their comment to apply here, and I’ll re-enable comments.

Organ epigenetic memory

This 2018 Japanese review subject was the relationships of organ memory and non-communicable diseases:

“Organ memory is the engraved phenotype of altered organ responsiveness acquired by a time-dependent accumulation of organ stress responses. This phenomenon is known as “metabolic memory” or “legacy effect,” which is similar to neuronal and immune memory.

Not only is the epigenetic change of key genes involved in the formation of organ memory but the alteration of multiple factors, including low molecular weight energy metabolites, immune mediators, and tissue structures, is involved as well. These factors intercommunicate during every stress response and carry out incessant remodeling in a certain direction in a spiral fashion through positive feedback mechanisms.

The systematic review revealed that each intervention type, that is:

  • Glucose lowering,
  • Blood pressure lowering, or
  • LDL-cholesterol lowering,

possessed unique characteristics of the memory phenomenon. Most of the observational periods of these studies lasted for > 10 years. Memory phenomenon was suggested to last for a long time and is thought to have a considerable effect on the clinical course of NCDs [non-communicable diseases].

Organs cannot possess consciousness, so it might not be appropriate to consider whether a recalling process exists in organs. However, the properties of organs are incessantly altered by external stimuli loaded on organs as if it is updating.

It is clinically important to investigate whether organ memory can be updated by our behaviors. Once organ memory is established in an organ, organ memory in each organ can influence one another and affect organ memory in a different organ.

Epigenome-modification enzymes, such as histone deacetylases and DNA methyltransferases, and transcription factors seem to be essential for the epigenetic regulation of gene expression, which is involved in the generation of organ memory. Cellular metabolism can epigenetically modulate the expression of genes that are related to the progression of diseases.”

1. The reviewers asserted:

“Organs cannot possess consciousness, so it might not be appropriate to consider whether a recalling process exists in organs.”

Memory studies don’t require this consciousness to investigate even the brain organ’s areas and functions. Researchers observe memory by measuring stimulus/response items like neuron activation and various levels of behavior. Consciousness is an emergent property.

2. Regarding recall: An organ’s “engraved phenotype of altered organ responsiveness” may not have recall itself, but it doesn’t have a separate existence apart from its body. An organ can’t be removed from its body for very long and still be part of its body.

When an organ is in its normal state as part of a body, it has access to recall-like functions via the “inter-organ communication of organ memory.” The review also mentioned:

“Organ memory in each organ can influence one another and affect organ memory in a different organ.

Evolution didn’t support unnecessary duplication for a kidney’s memory to include recall because it’s part of a body that includes a brain that has recall. Evolution didn’t duplicate functions of a kidney’s memory in a brain, either.

https://www.nature.com/articles/s41440-018-0081-x “Organ memory: a key principle for understanding the pathophysiology of hypertension and other non-communicable diseases” (not freely available)

Flawed epigenetic measurements of behavioral experiences

This 2018 New York rodent study not only wasted resources but also speciously attempted to extrapolate animal study findings to humans:

“While it is clear that behavioral experience modulates epigenetic profiles, it is less evident how the nature of that experience influences outcomes and whether epigenetic/genetic “biomarkers” could be extracted to classify different types of behavioral experience.

Male and female mice were subjected to either:

  • a Fixed Interval (FI) schedule of food reward, or
  • a single episode of forced swim followed by restraint stress, or
  • no explicit behavioral experience

after which global expression levels of two activating (H3K9ac and H3K4me3) and two repressive (H3K9me2 and H3k27me3) post-translational histone modifications (PTHMs), were measured in hippocampus (HIPP) and frontal cortex (FC).

A random subset of 5 of the 12 animals from each sex/behavioral experience group were used for these analyses. FC and HIPP were dissected from each of those 5 brains and homogenized for subsequent analyses. Thus, sample size for PTHM expression levels was n = 5 for each region/sex/behavioral treatment group and all PTHM expression level analyses utilized the homogenized tissue.

The specific nature of the behavioral experience differentiated profiles of PTHMs in a sex- and brain region-dependent manner, with all 4 PTHMs changing in parallel in response to different behavioral experiences. Global PTHMs may provide a higher-order pattern recognition function.”

The researchers knew or should have known that measuring “global expression levels” in “homogenized tissue” of “n = 5” subjects was flawed, and they did it anyway. They acknowledged some of the numerous study design defects with qualifiers such as:

“Even though these were global levels of histone modifications (and thus not indicative of changes at specific genes or sites on genes)..

As FS-RS behavioral experience was completed before FI behavioral experience, a longer overall post-behavior experience time (approximately 1 week) elapsed for this group, resulting in some differences in overall timing between these experiences and global PTHM assessment. However, extending the duration of the FS-RS experience (i.e., repeated exposures) would also have led to habituation..”

Did they purposely make these mistakes because of the “biomarkers” paradigm?

What would they have found if they had followed their judgments and training to design a better study? Experience-dependent histone modifications that differed by gender and brain region was certainly a promising research opportunity.

As for extrapolating the cited animal study findings to humans? Ummm..NO!

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6060276/ “Different Behavioral Experiences Produce Distinctive Parallel Changes in, and Correlate With, Frontal Cortex and Hippocampal Global Post-translational Histone Levels”

Unindexed comment links?

It’s dawned on me that although links in blog posts are indexed by search engines, links in comments may not be. Here’s a post to elevate links in three comments that may have escaped notice.

From A review of biological variability:

“It is my view that all researchers have a narrow focus on what they want to research, without having an over-riding paradigm in which to fit the research and its results. Janovian Primal Therapy and theory, with its focus and understanding of the three different levels of consciousness would provide for a much needed over-arching paradigm, especially in the area of mental health.”

Congratulations on an excellent podcast, Gil!
59. Gilbert Bates in “Feel It Still” // Love, Primal Therapy & the Three Levels of Consciousness

From Remembering Dr. Arthur Janov:

“You are right on. The Norcross survey, in particular, is utter crap. More than half of those “experts” surveyed were CBT therapists who knew nothing about PT and yet deemed themselves confident to judge “primal scream therapy” as “discredited.” I feel the therapy will never be understood for what it is.”

Thanks for the detailed explanation, Bruce!
The Worst Comparative Psychotherapy Study Ever Published

From How one person’s paradigms regarding stress and epigenetics impedes relevant research:

“There is of course, reversibility. Michael Meaney’s baby rats had their epigenetic changes reversed with loving maternal care. There are several compounds in development which have been shown to reverse methylation. This former physician and researcher says, “Epigenetic changes affect the level of activity of our genes. Genetic activity levels affect our emotions, beliefs, and our bodies. Exploring epigenetics and chronic illness may help us understand causes that many of us suspect have played a role in the onset and evolution of our illnesses. Furthermore, these epigenetic changes have been found to be reversible, at least some of the time, even with a seemingly indirect treatment such as psychotherapy.” Epigenetics and Chronic Illness: Why Symptoms May Be Reversible

I looked up the psychotherapy references and found this: Serotonin tranporter methylation and response to cognitive behaviour therapy in children with anxiety disorders (reversible even with CBT, the weakest therapy of all!)

And this:
MAOA gene hypomethylation in panic disorder—reversibility of an epigenetic risk pattern by psychotherapy (also CBT)

So what gives? I suspect that your researcher is working with his/her head in the sand, hamstrung by their ideological biases. If CBT can effect epigenetic changes, imagine what primal therapy can do.”

And a seven-year anniversary repost of events that affect me every day:

Reflections on my four-year anniversary of spine surgery

Prenatal programming of human HPA axis development

This 2017 UC Irvine human review subject provided details of how fetal hypothalamic-pituitary-adrenal components and systems develop, and how they are epigenetically changed by the mother’s environment:

“The developmental origins of disease or fetal programming model predicts that intrauterine exposures have life-long consequences for physical and psychological health. Prenatal programming of the fetal hypothalamic-pituitary-adrenal (HPA) axis is proposed as a primary mechanism by which early experiences are linked to later disease risk.

Development of the fetal HPA axis is determined by an intricately timed cascade of endocrine events during gestation and is regulated by an integrated maternal-placental-fetal steroidogenic unit. Mechanisms by which stress-induced elevations in hormones of maternal, fetal, or placental origin influence the structure and function of the emerging fetal HPA axis are discussed.

Human gestational physiology and fetal HPA axis development differ even from that of closely related nonhuman primates, thereby limiting the generalizability of animal models. This review will focus solely on studies of prenatal stress and fetal HPA axis development in humans.”

Every time I read a prenatal study I’m in awe of all that has to go right, and at the appropriate time, and in sequence, for a fetus to be undamaged. Add in what needs to happen at birth, during infancy, and throughout early childhood, and it seems impossible for any human to escape epigenetic damage.

1. The reviewers referenced human research performed with postnatal subjects, as well as animal studies, despite the disclaimer:

This review will focus solely on studies of prenatal stress and fetal HPA axis development in humans.”

This led to blurring of what had been studied or not with human fetuses regarding the subject.

2. The reviewers uncritically listed many dubious human studies that had both stated and undisclosed severe limitations on their findings. It’s more appropriate for reviewers to offer informed reviews of cited studies, as Sex-specific impacts of childhood trauma summarized with cortisol:

“Findings are dependent upon variance in extenuating factors, including but not limited to, different measurements of:

  • early adversity,
  • age of onset,
  • basal cortisol levels, as well as
  • trauma forms and subtypes, and
  • presence and severity of psychopathology symptomology.”

3. It would have been preferable had the researchers stayed with their stated intention and critically reviewed only a few dozen studies with solid evidence of the review title: “Developmental origins of the human hypothalamic-pituitary-adrenal axis.” Let other reviews cover older humans, animals, and questionable evidence.

I asked the reviewers to provide a searchable file so that their work could be better used as a reference.

https://www.researchgate.net/publication/318469661_Developmental_origins_of_the_human_hypothalamic-pituitary-adrenal_axis “Developmental origins of the human hypothalamic-pituitary-adrenal axis” (registration required)