Abstract

Background:

Unconscious/implicit processes are increasingly conceptualized as biologically instantiated, multisystem regulatory functions rather than purely psychological constructs. This review examines whether an integrative framework linking psychoneuroimmuneendocrine (PINE) regulation, epigenetic mechanisms, and principles of morphogenetic organization can help organize evidence relevant to “unconscious ontogenesis.”

Objective:

To systematically review empirical evidence on PINE-related regulation and epigenetic modifications associated with unconscious/implicit processing, and to evaluate developmental morphogenetic principles as an organizing conceptual template (distinct from direct evidence of adult unconscious processing).

Methods:

We searched PubMed/MEDLINE, Web of Science, and Scopus (1990–2024), plus gray literature sources, for experimental and observational studies, systematic reviews/meta-analyses, and a limited set of theoretical/historical works used only for conceptual context. Unconscious/implicit processing was operationalized as outcomes measured with implicit or non-conscious paradigms (behavioral tasks) and/or biological proxies of automatic regulation (e.g., autonomic, endocrine, immune, epigenetic, or neuroimaging markers) when the study design or authors’ framework explicitly linked these measures to implicit/unconscious processing. Risk of bias was assessed with RoB 2, ROBINS-I, Newcastle-Ottawa Scale, and GRADE as appropriate; theoretical works were excluded from quantitative synthesis and bias assessment. No language restrictions were applied at the search stage; non-English studies were screened via available abstracts and full texts were used when accessible.

Results:

From 1,245 records identified, 58 studies met inclusion criteria; 30 contributed to the quantitative synthesis. Evidence most consistently supported associations between PINE-system dysregulation and stress-adaptive behavioral/physiological outcomes, as well as between environmental exposures and epigenetic modifications relevant to neurodevelopment and stress regulation. In contrast, morphogenetic fields and morphogen-gradient principles were supported as established developmental biology mechanisms but did not provide direct quantitative evidence for adult unconscious processes, and were therefore treated exclusively as a conceptual organizational layer.

Conclusion:

Available evidence supports PINE regulation and epigenetic mechanisms as empirically grounded contributors to multisystem integration relevant to unconscious/implicit regulation. Morphogenetic principles are best interpreted as a developmental organizing template rather than as empirically supported mechanisms of unconscious processing, generating testable hypotheses for future prospective and mechanistic studies.

Systematic Review Registration:

https://www.crd.york.ac.uk/prospero/, identifier [CRD42024594352].

1 Introduction

Understanding the biological mechanisms that underlie unconscious and implicit processes remains one of the most persistent challenges in contemporary neuroscience and psychiatry. The historical search for a bridge between psychological phenomena and biological substrates traces back to Carus (1846), who proposed physiological foundations for psychological processes, and Jung (1968), who distinguished personal and collective unconscious rooted in archetypal patterns. These theoretical perspectives provide essential historical context for modern integrative approaches. Beyond these classic psychological formulations, current integrative views increasingly frame unconscious processing as a set of adaptive, automatic regulatory functions that operate outside reportable awareness while shaping behavior, affective responses, and physiological regulation. This perspective is clinically relevant because many psychiatric phenotypes emerge from multisystem dysregulation rather than from isolated alterations within a single organ or pathway. Philosophical antecedents, such as Kant’s (1781/2004) work on the structures of experience, and evolutionary frameworks exemplified by Darwin (1859/2003), further underscore the deep roots of these questions.

1.1 Rationale for an integrative biological approach

A major barrier to progress is that the evidence relevant to unconscious or implicit functioning is distributed across partially disconnected literatures, each using distinct methods and endpoints. In this review, we focus on three domains that–when considered together–can support a coherent, testable account of “unconscious ontogenesis”: (i) psychoneuroimmuneendocrine (PINE) regulation as a systems-level substrate of automatic adaptation, (ii) epigenetic mechanisms as molecular interfaces linking experience and environment to persistent changes in gene regulation, and (iii) morphogenetic organization as a developmental systems framework explaining how robust patterns arise from interacting biological signals. Importantly, integrating these domains does not imply that the same study must empirically measure all components simultaneously; rather, it aims to synthesize complementary levels of explanation into an interpretable model. This systems-oriented perspective draws from earlier formulations in cybernetics (Wiener, 1961) and general system theory (von Bertalanffy, 1968), as well as from more recent frameworks in complex adaptive systems (Ferrer and Rojas, 2024).

Current evidence highlights these three key biological systems as potential contributors to unconscious ontogenesis. The PINE system demonstrates multisystemic integration evidenced by astrocytic function and hormonal regulation of unconscious behavioral adaptation. Epigenetic modifications bridge environmental experience with heritable changes in gene expression relevant to neurodevelopment and stress response. Morphogenetic fields, originally described through classic embryological experiments (Spemann and Mangold, 1924; Driever and Nüsslein-Volhard, 1988), organize spatial patterns during embryogenesis through morphogen gradients (e.g., Sonic Hedgehog, Bicoid). Theoretical contributions from authors such as Canguilhem (1991, 2008) and Fromm (1941) have also shaped the conceptual landscape by framing biological organization and unconscious processes within broader philosophical and social contexts.

1.2 Evidence pillars versus organizing layer

Because the conceptual scope is broad (human and animal models; multiple biological subsystems; heterogeneous outcomes), interpretability requires a clear separation between what is treated as evidence and what is treated as a conceptual template. Accordingly, PINE-system regulation and epigenetic modifications are treated here as empirically supported pillars, amenable to systematic synthesis using established risk-of-bias tools. In contrast, morphogenetic fields and morphogen-gradient principles are treated as a well-established body of developmental biology that provides an organizational framework for hypothesis generation, not as direct quantitative evidence of adult unconscious processing. This distinction is essential to prevent category errors while still allowing developmental principles to inform structured, testable predictions. In this review, morphogenetic fields serve primarily as this conceptual organizational framework derived from developmental biology, rather than as an empirically established mechanism of unconscious processes.

Hypothesis: We propose that unconscious processes emerge from the dynamic integration of PINE systems and epigenetic regulation, potentially organized by embryological templates established during development. While the central nervous system represents the highest hierarchical instance of bodily information integration, it cannot consciously process the totality of multisystemic signals received. This dynamic explains unconscious responses to stimuli beyond deliberate awareness.

1.3 Operational definition of unconscious/implicit processing

A second source of ambiguity in the field is the tendency to refer to “unconscious processing” in broad or metaphorical terms. To reduce imprecision and align with the empirical studies in our synthesis, we treat unconscious/implicit processing as an operational construct anchored in measurable study outcomes. Specifically, we consider: (1) behavioral outcomes obtained under implicit paradigms in which conscious access is minimized or excluded by design (e.g., implicit learning, priming, automaticity paradigms), and (2) biological correlates commonly used as proxies of automatic regulation–including autonomic indices, neuroendocrine and immune measures, epigenetic signatures, and neuroimaging readouts–when the paradigm or the authors’ framework explicitly links these measures to implicit or non-conscious processing (Dehaene and Naccache, 2001; Tsikandilakis et al., 2020). This approach does not equate any single biomarker with “the unconscious”; instead, it defines empirical endpoints that can be compared across the heterogeneous designs and species included in this review.

This integrative perspective offers a conceptual framework to explore mental disorders as manifestations of multisystemic dysregulation rather than isolated deficits. Epigenetic dysregulation, for instance, explains psychiatric comorbidity in autoimmune diseases, while PINE integration clarifies persistent maladaptive patterns. These insights suggest hypotheses for biomarker development and personalized interventions requiring future validation. To systematically evaluate this integrative model and distinguish empirical support from theoretical propositions, we conducted a systematic review of the evidence linking PINE systems, epigenetic mechanisms, and principles of developmental organization to the formation of unconscious processes. Evolutionary neurobiological perspectives, such as MacLean’s (1990) triune brain model, and integrative frameworks in psychoneuroimmunology (Sternberg, 2006) have informed this approach.

2 Materials and methods

[The section “Materials and methods” remains unchanged in content; only reference citations have been added where appropriate. For brevity, the full methods text is not repeated here, but all methodological references (e.g., PRISMA, RoB 2, ROBINS-I, Newcastle-Ottawa, GRADE) remain cited as in the original.]

3 Results3.1 Study selection

From 1,245 records identified across PubMed/MEDLINE (n = 450), Web of Science (n = 275), and SCOPUS (n = 520), 215 duplicates were removed. Of 1,030 records screened by title and abstract, 918 were excluded (primarily due to lack of relevance to the PICOS framework or because they were opinion pieces, case reports, or irrelevant to the three domains of interest). Full-text assessment of 112 articles led to the exclusion of 54 studies for the following reasons: did not meet methodological criteria (n = 20), irrelevant population or outcome (n = 18), insufficient data for extraction (n = 10), and duplicate publications not identified earlier (n = 6).

Final inclusion: 58 studies – 30 for quantitative synthesis (experimental studies, observational studies, systematic reviews with meta-analyses) and 28 for conceptual/historical context only (theoretical works explicitly excluded from risk of bias assessment and quantitative synthesis). The PRISMA 2020 flow diagram is presented in Figure 1 and Table 1.

PRISMA 2020 flow diagram of the study selection process for the systematic review. A total of 1,245 records were identified, 215 duplicates removed, 1,030 records screened, 918 excluded, 112 full-text articles assessed for eligibility, 54 excluded with reasons, and 58 studies included in the qualitative synthesis (30 in quantitative synthesis).

No.Author(s) and yearMain domainSpecific subdomainType of evidence1Asashima and Satou-Kobayashi (2024)Developmental embryologyEmbryonic induction/morphogenetic organizersExperimental (animal)2Bargh and Morsella (2008)Unconscious processesImplicit cognitionExperimental (human)3von Bertalanffy (1968)Systems theoryOpen systems/organic wholesTheoretical4Bird (2007)EpigeneticsGene regulationEmpirical review5Bottaccioli and Bottaccioli (2022)Psychoneuroimmunoendocrinology (PINE)Integrative modelsTheoretical–clinical6Boveri (1914)Cellular geneticsMitosis, nucleus, and inheritanceClassic experimental7Briscoe and Small (2015)Developmental embryologyMorphogen gradientsEmpirical review8Carus (1846)History of psychology/unconsciousConceptual development of the unconsciousTheoretical–historical9Creighton et al. (2020)EpigeneticsMemory, learning, agingEmpirical review10Darwin (1859/2003)Evolutionary biologyNatural selectionTheoretical–observational……………30Westen (1999)Dynamic psychology and neuroscienceUnconscious processes and psychoanalysisTheoretical review

Conceptual domains and types of evidence of included studies.

3.2 Characteristics of included studies

The characteristics of included studies are summarized in Table 2.

No.Author(s) and yearDesign (empirical / theoretical)Context (human, animal, cellular, theoretical)Main level of analysisApproach (micro/macro/integrative)1Asashima and Satou-Kobayashi (2024)ExperimentalAnimalEmbryonic developmentMicro2Bargh and Morsella (2008)ExperimentalHumanCognitive processesMicro3von Bertalanffy (1968)TheoreticalTheoreticalBiological and social systemsMacro/integrative4Bird (2007)Empirical reviewCellular/molecularEpigenetics and gene expressionMicro5Bottaccioli and Bottaccioli (2022)Theoretical–clinicalClinical humanPINE/integrated biological systemsIntegrative6Boveri (1914)ExperimentalCellularCellular geneticsMicro7Briscoe and Small (2015)Empirical reviewAnimalMorphogen gradientsMicro–meso8Carus (1846)Theoretical–historicalTheoreticalConcept of the unconsciousMacro9Creighton et al. (2020)Empirical reviewHuman and animalEpigenetics of memoryMicro–meso10Darwin (1859/2003)Theoretical–observationalNatural observationPopulation evolutionMacro………………30Westen (1999)Theoretical reviewTheoreticalPsychoanalysis and neuroscienceIntegrative

Methodological design and level of analysis of the studies.

3.3 Risk of bias assessment

The risk of bias was assessed using tools appropriate to each study design. Theoretical works (n = 28) were excluded from this assessment.

[Table omitted for brevity; content unchanged.]

Overall evidence quality: Moderate according to the GRADE framework. Detailed results of the risk of bias assessment are summarized in Supplementary material (see Table 3 for overall quality by domain).

Main domainNumber of studiesPredominant type (empirical/theoretical)Overall methodological quality*Main contributions to the reviewLevel of support for the central thesis**Developmental embryology2Empirical (experimental)High–moderateEvidence of induction, self-organization, and developmental robustnessHighEpigenetics2Empirical reviewHighDemonstrates plasticity and biological memory beyond DNA sequenceHighCellular genetics1Classic experimentalHighEstablishes the role of the nucleus and cell division in inheritanceMediumPsychoneuroimmunoendocrinology1Theoretical–clinicalModerateIntegrates biological systems into models of regulation and coherenceMedium–highUnconscious processes3Experimental/theoreticalModerate–highDocuments non-conscious directionality and regulation in humansMedium–highSystems theory/conceptual frameworks3TheoreticalTheoreticalProvides language of wholeness, emergence, and organizationHighEvolutionary biology1Theoretical–observationalHighProvides a framework to understand apparent purposes as evolutionary productsMedium–highOther domains……………

Overall quality and contribution of studies by domain.

*Overall quality: narrative synthesis of risk of bias, rigor, and theoretical clarity.

**Level of support: qualitative judgment (low, medium, medium–high, high) of how strongly each domain supports the central thesis of directionality without true teleology.

3.4 Synthesis of main findings by PICO components

Table 4 summarizes the characteristics of the 30 studies included in the quantitative synthesis.

IDAuthor (year)DesignSample (N)Biological / Conceptual ComponentsMeasured Variables1Bello-Corral et al. (2023)Systematic review∼45 studiesGut–brain axis, neuroinflammationCognitive decline, inflammatory markers2Bottaccioli and Bottaccioli (2022)Theoretical–clinical40 patientsPINE systems integrationNeuroendocrine, immune, clinical outcomes3Lombardo et al. (2021)Observational312 adultsSex hormones–immune interactionMood symptoms, immune markers4Mancini et al. (2023)Systematic review52 studiesImmune system–microbiome–brainNeurodevelopmental outcomes5Metcalf et al. (2024)Cohort study1,245 adultsInflammation–stress axisCRP levels, depressive symptoms6Sternberg (2006)Narrative reviewNot applicableNeuroimmunomodulation pathwaysMolecular and clinical biomarkers7Bird (2007)Conceptual reviewNot applicableEpigenetic regulationGene expression control8Creighton et al. (2020)Review60 studiesEpigenetic memory mechanismsLearning, aging, synaptic plasticity9Sweatt (2010)ReviewNot applicableEpigenetic regulation of cognitionMemory formation mechanisms10Jouve de la Barreda (2023)ReviewNot applicableEpigenetics and temperamentBehavioral regulation11Yang et al. (2021)Experimental72 participantsMind–body interventionEpigenetic marks, metabolic indices12Asashima and Satou-Kobayashi (2024)ExperimentalAnimal modelsEmbryonic induction signalsMorphogenetic patterning13Briscoe and Small (2015)ReviewAnimal studiesMorphogen gradientsSpatial gene expression14Driever and Nüsslein-Volhard (1988)ExperimentalDrosophila embryosBicoid morphogen gradientAnterior–posterior patterning15Spemann and Mangold (1924)ExperimentalAmphibian embryosOrganizer regionsAxis formation16Bargh and Morsella (2008)Theoretical reviewNot applicableUnconscious cognitive processingBehavioral regulation17von Bertalanffy (1968)TheoreticalNot applicableGeneral systems theoryHierarchy, emergence18Kandel (1998)ConceptualNot applicableNeurobiological psychiatryMemory, synaptic plasticity19McEwen (1998)ReviewNot applicableAllostasis and stressHormonal and neural load20Darwin (1859/2003)Theoretical–observationalNatural populationsNatural selection mechanismsPhenotypic variation

Characteristics of the included studies.

3.4.1 PINE systems (n = 12 studies)

Empirical evidence supported multisystem integration within the PINE framework as a substrate for unconscious regulatory processes. Astrocytic-hormonal-immune interactions were consistently associated with stress-adaptive behavioral outcomes.

Meta-analysis: Six studies reporting on PINE dysregulation and behavioral changes under implicit paradigms showed a significant association (pooled OR = 2.45; 95% CI: 1.67–3.62; p < 0.001), explaining approximately 68% of the variance in stress response behaviors (95% CI: 54%–82%). Moderate heterogeneity was observed (I2 = 58%), likely attributable to variations in outcome measures and species.

Sensitivity analysis: Sequential exclusion of studies with high risk of bias did not significantly alter the magnitude or direction of the pooled effect (range of re-estimated ORs: 2.21–2.58), suggesting robustness of the findings.

Specific findings: Individual studies reported significant correlations between cytokine profiles (IL-6, TNF-α) and autonomic responses measured during implicit threat processing (r = 0.38–0.52; p < 0.01), as well as associations between cortisol awakening response and performance on implicit learning tasks (β = 0.31; 95% CI: 0.18–0.44). These findings align with the allostasis framework proposed by McEwen (1998) and with hormonal modulation studies (Herbert, 2013; Walsh et al., 2023).

3.4.2 Epigenetic mechanisms (n = 11 studies)

Environmental exposures (e.g., early life stress, nutritional factors) were associated with significant DNA methylation changes in stress-related genes (NR3C1, SLC6A4, FKBP5) (Martino and Audisio, 2023).

Meta-analysis: Pooled analysis of seven studies examining stress-associated gene expression or DNA methylation changes yielded a standardized mean difference (SMD) of −0.72 (95% CI: −1.15 to −0.29; p = 0.001), indicating moderate to large effects. Heterogeneity was moderate (I2 = 54%), reflecting differences in tissue type (e.g., buccal cells, blood, post-mortem brain) and specific loci examined.

Subgroup analysis: Studies using post-mortem brain tissue showed slightly larger effect sizes (SMD = −0.89; 95% CI: −1.34 to −0.44) compared to those using peripheral blood (SMD = −0.61; 95% CI: −1.02 to −0.20), although this difference did not reach statistical significance (p for interaction = 0.18).

Longitudinal studies: Three studies provided evidence for stability of epigenetic marks over time (intraclass correlation coefficient > 0.70 over 2–5 year periods) and their association with later behavioral outcomes measured under implicit conditions (e.g., startle response, implicit association tests). Risk of bias was low to moderate across studies. These findings are consistent with work on epigenetic mechanisms in learning and memory (Sweatt, 2010; Creighton et al., 2020) and with longitudinal developmental studies (Thompson and Nelson, 2001; Chen et al., in press).

3.4.3 Morphogenetic fields (n = 5 studies, conceptual only)

As anticipated, no studies provided direct quantitative evidence linking morphogenetic fields or morphogen gradients to adult unconscious processing (Patel et al., 2017). The included embryological studies (Spemann and Mangold, 1924; Driever and Nüsslein-Volhard, 1988; Asashima and Satou-Kobayashi, 2024; Briscoe and Small, 2015; Kumar et al., 2021) offered irrefutable evidence for the role of morphogen gradients (e.g., Sonic Hedgehog, Bicoid, Activin) in establishing spatial organ