Impact of isolation method on cellular activation and presence of specific tendon cell subpopulations during in vitro culture.

Tendon injuries are common and heal poorly, due in part to a lack of understanding of fundamental tendon cell biology. A major impediment to the study of tendon cells is the absence of robust, well-characterized in vitro models. Unlike other tissue systems, current tendon cell models do not account for how differences in isolation methodology may affect the activation state of tendon cells or the presence of various tendon cell subpopulations. The objective of this study was to characterize how common isolation methods affect the behavior, fate, and lineage composition of tendon cell cultures. Tendon cells isolated by explant exhibited reduced proliferative capacity, decreased expression of tendon marker genes, and increased expression of genes associated with fibroblast activation compared to digested cells. Consistently, explanted cells also displayed an increased propensity to differentiate to myofibroblasts compared to digested cells. Explanted cultures from multiple different tendons were substantially enriched for the presence of scleraxis-lineage (Scx-lin+) cells compared to digested cultures, while the overall percentage of S100a4-lineage (S100a4-lin+) cells was dependent on both isolation method and tendon of origin. Neither isolation methods preserved the ratios of Scx-lin+ or S100a4-lin+ to non-lineage cells seen in tendons in vivo. Combined, these data indicate that further refinement of in vitro cultures models is required in order to more accurately understand the effects of various stimuli on tendon cell behavior. Statement of clinical significance: The development of informed in vitro tendon cell models will facilitate enhanced screening of potential therapeutic candidates to improve tendon healing.

Contrast effects in backward evaluative conditioning: Exploring effects of affective relief/disappointment versus instructional information.

Past studies of backward evaluative conditioning (EC) have found an assimilation effect, in that neutral conditional stimuli (conditional stimulus [CS]) were found to acquire the valence of co-occurring unconditional stimuli (US). Recent studies employing a concurrent forward and backward conditioning paradigm with instructions suggesting a contrastive relation between the US and the backward CS have resulted in contrast effects, in that backward CSs acquired valence opposite to the US. The current research investigated whether these effects were in fact due to the instructions highlighting the contrastive relation between the US and CS, or whether affective relief/disappointment experienced at US offset could account for this result. Consistent with the hypothesized role of instructions, backward CS contrast effects occurred only when instructions highlighted the valence of the US and attributed control of that US to the CSs. In contrast to the affective relief/disappointment hypothesis, no backward CS contrast effects were found without such instructions. (PsycInfo Database Record (c) 2021 APA, all rights reserved).

Startle during backward evaluative conditioning is not modulated by instructions.

Instructions highlighting that backward conditional stimuli (CSs) stop unconditional stimuli (USs) result in their acquiring valence opposite to that of the US on explicit measures of valence. We assessed whether such instructions would influence startle blink modulation in the same way. Two groups were presented with concurrent forward and backward evaluative conditioning (CS-US-CS) using cartoon aliens as CSs, and pleasant, neutral, and unpleasant sounds as USs. Startle magnitude was measured during conditioning and valence ratings were assessed after conditioning. Participants in the "start-stop" instructions group (n = 41) were instructed to learn whether CSs started or stopped US presentations, while participants in the "observe" instructions group (n = 41) were told to pay attention to the stimuli as they would be asked questions about them after the experiment. In the start-stop instructions group backward CSs paired with positive USs were rated as less pleasant than backward CSs paired with neutral and negative USs (contrast effect), whereas ratings of backward CSs did not differ in the observe instructions group. Startle magnitude was larger during backward CSs paired with positive USs in comparison to CSs paired with neutral or negative USs in both instruction groups. Startle blink modulation was unaffected by instructions, suggesting that startle indexes the emotional state at the time of probe presentation rather than CS valence based on propositional information about the function of the CS.

How disappointing: Startle modulation reveals conditional stimuli presented after pleasant unconditional stimuli acquire negative valence.

Past research on backward conditioning in evaluative and fear conditioning yielded inconsistent results in that self-report measures suggest that the conditional stimulus (CS) acquired the valence of the unconditional stimulus (US) in fear conditioning (assimilation effects), but the opposite valence in evaluative conditioning (contrast effects). Conversely, implicit measures of CS valence suggest assimilation effects in evaluative backward conditioning, whereas startle modulation indicates contrast effects in backward fear conditioning. The current study investigated whether US intensity could account for the dissociation on implicit measures between fear and evaluative conditioning. Self-report measures of evaluative learning indicated assimilation effects for forward conditioning, whereas backward contrast effects were observed with intense USs only. Blink startle modulation indicated assimilation effects in forward conditioning and contrast effects in backward conditioning, regardless of US intensity. Experiment 2 included a neutral US in order to assess whether the offset of the positive US elicits an opponent emotional response that mirrors relief (disappointment), which is thought to mediate the reduction in startle seen during backward CSs in fear conditioning. This opponent emotional response was evident as startle magnitude during backward CSs increased linearly with increasing US pleasantness. Omission of the forward CSs led to an assimilation effect in self-report measures. The current results extend our understanding of emotional learning to stimuli encountered after salient emotional events. Startle reflects the emotion prevailing after US offset, relief or disappointment, whereas self-report measures seem more attuned to factors such as US predictability and intensity.

Temporal context cues in human fear conditioning: Unreinforced conditional stimuli can segment learning into distinct temporal contexts and drive fear responding.

In associative learning, if stimulus A is presented in the same temporal context as the conditional stimulus (CS) - outcome association (but not in a way that allows an A-CS association to form) it becomes a temporal context cue, acquiring the ability to activate this context and retrieve the CS-outcome association. We examined whether a CS- presented during acquisition or extinction that predicted the absence of the unconditional stimulus (US) could act as a temporal context cue, reducing or enhancing responding, in differential fear conditioning. Two groups received acquisition (CSx-US, CSa-noUS) in phase 1 and extinction (CSx-noUS; CSe-noUS) in phase 2 (AE groups), and two groups received extinction in phase 1 and acquisition in phase 2 (EA groups). After a delay, participants were presented with either CSa (AEa and EAa groups) or CSe (AEe and EAe groups). Responding to CSx was enhanced after presentation of CSa but reduced after presentation of CSe, suggesting that training was segmented into two learning episodes and that the unreinforced CS present during an episode retrieved the CSx-US or CSx-noUS association. These findings suggest that temporal context cues may enhance or reduce fear responding, providing an exciting new avenue for relapse prevention research.