When pulmonary surfactant was first detected in the 1950s by Pattle and Clements, many thousands of infants perished each year due to a respiratory illness termed hyaline membrane disease. Hyaline membranes are formed by plasma leaking through damaged endothelial barriers into the terminal bronchiolar: alveolar spaces. Since the leaking plasma lacks erythrocytes, these clots are opaque. Insightful research by Avery and Mead soon led to the suggestion that the neonatal respiratory distress syndrome (RDS) did not arise because of the presence of hyaline membranes, but rather was related to the lack of sufficient pulmonary surfactant, mainly as a result of immaturity. Unfortunately, initial attempts at treating RDS with aerosolized dipalmitoyl-phosphatidylcholine, the major single molecular component, proved unsuccessful. Almost 20 years later, it was demonstrated by Enhorning and Robertson that treating prematurely delivered rabbit pups with natural surfactant prevents respiratory failure. Initially, it appeared unlikely that animal surfactants could be used for therapy with human infants. However, in 1980, Fujiwara demonstrated that a modified bovine surfactant extract promoted gaseous exchange with infants suffering from RDS. Soon a number of bovine and porcine-modified surfactants and two wholly synthetic formulations were shown to alleviate RDS. The present review relates some of the key scientific findings and significant clinical contributions responsible for reducing the neonatal morbidity and mortality associated with RDS. It further describes some of the more recent findings on the biological, biophysical, and physiological significance of pulmonary surfactant in health and disease.

Pulmonary surfactant is a critical component of lung function in healthy individuals. It functions in part by lowering surface tension in the alveoli, thereby allowing for breathing with minimal effort. The prevailing thinking is that low surface tension is attained by a compression-driven squeeze-out of unsaturated phospholipids during exhalation, forming a film enriched in saturated phospholipids that achieves surface tensions close to zero. A thorough review of past and recent literature suggests that the compression-driven squeeze-out mechanism may be erroneous. Here, we posit that a surfactant film enriched in saturated lipids is formed shortly after birth by an adsorption-driven sorting process and that its composition does not change during normal breathing. We provide biophysical evidence for the rapid formation of an enriched film at high surfactant concentrations, facilitated by adsorption structures containing hydrophobic surfactant proteins. We examine biophysical evidence for and against the compression-driven squeeze-out mechanism and propose a new model for surfactant function. The proposed model is tested against existing physiological and pathophysiological evidence in neonatal and adult lungs, leading to ideas for biophysical research, that should be addressed to establish the physiological relevance of this new perspective on the function of the mighty thin film that surfactant provides.

Introduction The dramatic impact of COVID-19 on humans worldwide has initiated an extraordinary search for effective treatment approaches. One of these is the administration of exogenous surfactant, which is being tested in ongoing clinical trials. Areas covered Exogenous surfactant is a life-saving treatment for premature infants with neonatal respiratory distress syndrome. This treatment has also been tested for acute respiratory distress syndrome (ARDS) with limited success possibly due to the complexity of that syndrome. The 60-year history of successes and failures associated with surfactant therapy distinguishes it from many other treatments currently being tested for COVID-19 and provides the opportunity to discuss the factors that may influence the success of this therapy. Expert opinion Clinical data provide a strong rationale for using exogenous surfactant in COVID-19 patients. Success of this therapy may be influenced by the mechanical ventilation strategy, the timing of treatment, the doses delivered, the method of delivery and the preparations utilized. In addition, future development of enhanced preparations may improve this treatment approach. Overall, results from ongoing trials may not only provide data to indicate if this therapy is effective for COVID-19 patients, but also lead to further scientific understanding and improved treatment strategies.

Cholinephosphotransferase, the enzyme responsible for the terminal reaction in the formation of phosphatidylcholine (PC) in lung, has been extensively examined in lung. Pulmonary surfactant stabilizes the lung by reducing the surface tension at the air-liquid interface of the alveolar surface. The deacylation-reacylation, or "Land's cycle", can account for many of the observations made with fractions from whole lung or type II cells. The crucial role of cholinephosphate cytidylyltransferase in the increase in PC production is further suggested by studies with fetal rat lung. The disaturated PC in lung can be synthesized partly via the de novo pathway, but also through a progressive conversion of unsaturated PCs generated by the de novo pathway into their disaturated counterparts. The lack of correlation between the increase in the incorporation of choline into PC and the activities of the enzymes involved in PC production prompted examination of the pool sizes of choline and its intermediates in rabbit lung.

BackgroundThe complex branching structure of the lung makes direct delivery of drugs, such as the antimicrobial peptides cathelicidins, to the small and distal airways a major challenge. To address this issue our lab has been investigating the use of exogenous surfactant (BLES), to facilitate the pulmonary delivery of cathelicidins. BLES is a lipoprotein complex capable of spreading through the lung. Its major lipid components are phosphatidylglycerol (PG; 10%), phosphatidylcholine (PC; 40%) and dipalmitoylphosphatidylcholine (DPPC;40%). Although, BLES in combination with cathelicidins has shown promising results, the presence of BLES has also been shown to significantly reduce the ability of these peptides to kill bacteria.Objectives1) Determine the lipid components of BLES that are limiting the bactericidal effects of cathelicidins, 2) reconstitute BLES to reduce its inhibitory effect on the antimicrobial properties of cathelicidins, and 3) assess the spreading and bacterial killing of reformulated BLES/cathelicidin preparations at a distal site.HypothesisA reformulated BLES will improve the bacterial killing of BLES/cathelicidin mixtures, without reducing spreading to distal sites in vitro.MethodsAntimicrobial assays were performed against a lab strain of Pseudomonas aeruginosa for four cathelicidins in the presence of individual surfactant lipids. BLES was also supplemented with different lipids to generate different relative amounts of PG (5–40%), PC (20–70%) and DPPC (20–70%). These different versions of BLES were combined with cathelicidins for bacterial killing curves and assessment on our newly developed wet bridge transfer system. This in vitro system was utilized to assess spreading and bacterial killing at distal sites. P. aeruginosa was seeded to the distal well and cathelicidins alone or resuspended in a reformulated BLES were administered to the delivery well.ResultsAt concentrations found in BLES, PG was the only lipid component to significantly inhibit the bacterial killing of all cathelicidins tested. Furthermore, all reformulated surfactant preparations with lower PG concentrations than BLES were shown to significantly enhance peptide function compared to BLES. Experiments performed on the wet bridge transfer system showed that altering BLES to limit PG content (6–9%) significantly enhanced bacterial killing in the distal dish for suspended cathelicidins compared to BLES/cathelicidin. However, reducing the PG content of BLES to 5% was shown to significantly inhibit its ability to spread across the wet bridge transfer system. Moreover, when combined with cathelicidins it was shown have significantly lower bacterial killing in the distal dish compared to BLES.DiscussionThese results suggest that PG is the major component of BLES inhibiting the bacterial killing properties of cathelicidins. Furthermore, the data from the wet bridge experiments indicated that altering the lipid composition of BLES to limit PG‐content creates a more effective cathelicidin/surfactant preparation. Therefore, future studies should utilize artificial surfactants with lower levels of PG to delivery cathelicidins to the peripheral airways. However, some PG may be needed for peptide transfer and normal surfactant function.Support or Funding InformationLawson Internal Research FundThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Due to its branching structure, drug delivery to the peripheral areas of the lung is a major challenge. Consequently, most pulmonary therapies utilize large systemic dosing, with the potential for adverse side effects. One proposed strategy to overcome this challenge is to use exogenous surfactant, a material capable of distributing throughout the lung, as a pulmonary drug delivery vehicle. The objective was to develop and test an in vitro system to rapidly assess surfactant based therapies prior to animal studies. The Wet Bridge Transfer System consisted of two connected wells in which drugs were instilled into a delivery well and function was tested in a remote well which mimicked the remote areas of the lung where drug activity would be required. The system was used to assess surfactant as a carrier for antibiotics (Gentamicin, Ciprofloxacin, and Colistin) by measuring their ability to kill Pseudomonas aeruginosa bacteria in the remote well. Anti-inflammatory agents (Budesonide and a host defense peptide, CATH-2) with and without exogenous surfactant were examined using stimulated macrophages in the remote well and IL-6 concentration as an outcome. The results showed that being paired with surfactant, Gentamicin and Ciprofloxacin, but not Colistin, had significantly greater bacterial killing in the remote wells. Similarly, when combined with a surfactant, both Budesonide and CATH-2 significantly lowered IL-6 concentrations. We conclude that the wet-bridge system can be used to rapidly screen surfactant-based therapies prior to their assessment in vivo. Furthermore, exogenous surfactant was an effective delivery vehicle for several antimicrobial and anti-inflammatory therapeutics.

Human cysteine cathepsins (Cats) are implicated in lung injuries and tissue remodeling and have recently emerged as important players in pulmonary inflammations. The proteolytic activities of Cat B, L, K, S and H are dramatically increased in the sputum of patients with cystic fibrosis (CF), suggesting a possible involvement in the CF pathophysiology. We found that pulmonary surfactant protein A (SP-A) that participates to innate host defense is extensively degraded in CF expectorations. Breakdown of SP-A was markedly decreased in CF sputum by E-64 and Mu-Leu-Hph-VSPh, a Cat S inhibitor. Cat S cleaved efficiently and specifically SP-A within critical residues of the solvent-exposed loop of its carbohydrate recognition (C-type lectin) domain that allows binding to pathogens. Cat S decreased aggregation properties of SP-A (self-aggregation, aggregation of phospholipid vesicles and rough LPS). Moreover cleavage of SP-A by Cat S reduced binding to yeast mannan and impaired agglutination of Escherichia coli and Pseudomonas aeruginosa, a foremost detrimental pathogen colonizing the lungs of CF patients. Besides human neutrophil serine proteases and bacterial proteases, we propose that Cat S may participate in the pathophysiology of CF by weakening the antibacterial activity of SP-A. More broadly, present results provide further indication that Cat S, along with Cats B and L, could display immuno-modulatory functions by inactivating key proteins involved in the innate immunity defense.

Pulmonary surfactant lines the entire alveolar surface, serving primarily to reduce the surface tension at the air-liquid interface. Surfactant films adsorb as a monolayer interspersed with multilayers with surfactant lipids segregating into different phases or domains. Temperature variation, which influences lipid physical properties, affects both the lipid phase segregation and the surface activity of surfactants. In hibernating animals, such as 13-lined ground squirrels, which vary their body temperature, surfactant must be functional over a wide range of temperatures. We hypothesised that surfactant from the 13-lined ground squirrel, Ictidomys tridecemlineatus, would undergo appropriate lipid structural re-arrangements at air-water interfaces to generate phase separation, sufficient to attain the low surface tensions required to remain stable at both low and high body temperatures. Here, we examined pressure-area isotherms at 10, 25 and 37°C and found that surfactant films from both hibernating and summer-active squirrels reached their highest surface pressure on the Wilhelmy-Langmuir balance at 10°C. Epifluorescence microscopy demonstrated that films of hibernating squirrel surfactant display different lipid micro-domain organisation characteristics than surfactant from summer-active squirrels. These differences were also reflected at the nanoscale as determined by atomic force microscopy. Such re-arrangement of lipid domains in the relatively more fluid surfactant films of hibernating squirrels may contribute to overcoming collapse pressures and support low surface tension during the normal breathing cycle at low body temperatures.

The exact mechanism by which pulmonary surfactant films reach the very low surface tensions required to stabilize the alveoli at end expiration remains uncertain. We utilized the nanoscale sensitivity of atomic force microscopy (AFM) to examine phospholipid (PL) phase transition and multilayer formation for two Langmuir–Blodgett (LB) systems: a simple 3 PL surfactant-like mixture and the more complex bovine lipid extract surfactant (BLES). AFM height images demonstrated that both systems develop two types of liquid condensed (LC) domains (micro- and nano-sized) within a liquid expanded phase (LE). The 3 PL mixture failed to form significant multilayers at high surface pressure (π while BLES forms an extensive network of multilayer structures containing up to three bilayers. A close examination of the progression of multilayer formation reveals that multilayers start to form at the edge of the solid-like LC domains and also in the fluid-like LE phase. We used the elemental analysis capability of time-of-flight secondary ion mass spectrometry (ToF-SIMS) to show that multilayer structures are enriched in unsaturated PLs while the saturated PLs are concentrated in the remaining interfacial monolayer. This supports a modified squeeze-out model where film compression results in the hydrophobic surfactant protein-dependent formation of unsaturated PL-rich multilayers which remain functionally associated with a monolayer enriched in disaturated PL species. This allows the surface film to attain low surface tensions during compression and maintain values near equilibrium during expansion.

Gastrointestinal symptoms and altered blood phospholipid profiles have been reported in patients with autism spectrum disorders (ASD). Most of the phospholipid analyses have been conducted on the fatty acid composition of isolated phospholipid classes following hydrolysis. A paucity of information exists on how the intact phospholipid molecular species are altered in ASD. We applied ESI/MS to determine how brain and blood intact phospholipid species were altered during the induction of ASD-like behaviors in rats following intraventricular infusions with the enteric bacterial metabolite propionic acid. Animals were infused daily for 8 days, locomotor activity assessed, and animals killed during the induced behaviors. Propionic acid infusions increased locomotor activity. Lipid analysis revealed treatment altered 21 brain and 30 blood phospholipid molecular species. Notable alterations were observed in the composition of brain SM, diacyl mono and polyunsaturated PC, PI, PS, PE, and plasmalogen PC and PE molecular species. These alterations suggest that the propionic acid rat model is a useful tool to study aberrations in lipid metabolism known to affect membrane fluidity, peroxisomal function, gap junction coupling capacity, signaling, and neuroinflammation, all of which may be associated with the pathogenesis of ASD.

Hydrolysis of surfactant phospholipids (PL) by secretory phospholipases A(2) (sPLA(2)) contributes to surfactant damage in inflammatory airway diseases such as acute lung injury/acute respiratory distress syndrome. We and others have reported that each sPLA(2) exhibits specificity in hydrolyzing different PLs in pulmonary surfactant and that the presence of hydrophilic surfactant protein A (SP-A) alters sPLA(2)-mediated hydrolysis. This report tests the hypothesis that hydrophobic SP-B also inhibits sPLA(2)-mediated surfactant hydrolysis. Three surfactant preparations were used containing varied amounts of SP-B and radiolabeled tracers of phosphatidylcholine (PC) or phosphatidylglycerol (PG): 1) washed ovine surfactant (OS) (pre- and postorganic extraction) compared with Survanta (protein poor), 2) Survanta supplemented with purified bovine SP-B (1-5%, wt/wt), and 3) a mixture of dipalmitoylphosphatidylcholine (DPPC), 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC), and 1-palmitoyl-2-oleoyl-phosphatidylglycerol (POPG) (DPPC:POPC:POPG, 40:40:20) prepared as vesicles and monomolecular films in the presence or absence of SP-B. Hydrolysis of PG and PC by Group IB sPLA(2) (PLA2G1A) was significantly lower in the extracted OS, which contains SP-B, compared with Survanta (P = 0.005), which is SP-B poor. Hydrolysis of PG and PC in nonextracted OS, which contains all SPs, was lower than both Survanta and extracted OS. When Survanta was supplemented with 1% SP-B, PG and PC hydrolysis by PLA2G1B was significantly lower (P < 0.001) than in Survanta alone. When supplemented into pure lipid vesicles and monomolecular films composed of PG and PC mixtures, SP-B also inhibited hydrolysis by both PLA2G1B and Group IIA sPLA2 (PLA2G2A). In films, PLA2G1B hydrolyzed surfactant PL monolayers at surface pressures ≤30 mN/m (P < 0.01), and SP-B lowered the surface pressure range at which hydrolysis can occur. These results suggest the hydrophobic SP, SP-B, protects alveolar surfactant PL from hydrolysis mediated by multiple sPLA(2) in both vesicles (alveolar subphase) and monomolecular films (air-liquid interface).

Pulmonary surfactant stabilizes the lung by reducing surface tension (ST) to low values near zero at end expiration. Atomic force microscopy studies revealed that, as surface pressure increases, spread surfactant films initially form solid micro- and nanodomains and then generate 3-5 multilayers which apparently remain associated with the surface monolayer. Time-of-flight Secondary Ion Mass Spectroscopy analyses revealed selective squeeze-out must occur because the multilayers are enriched in unsaturated fluid phospholipids (PL) while the remaining monolayer becomes enriched in disaturated PL. Taken together, these results are consistent with a modified squeeze-out model where surfactant vesicles interact with the air-water interface through surfactant proteins B- and C-containing adsorption/fusion pores. PL migration onto the surface results in vesicle instability generating a monolayer at equilibrium surface pressure, which remains functionally associated with excess bilayer material. Initially, the monolayer and associated reservoir have similar composition, but film compression causes fluid PL to migrate through the fusion pores into the multilayers. Gel phase PL are restricted because they are sequestered in micro- or nanodomains. This process results in monolayers highly enriched in disaturated PL which reduce ST to near zero. Film expansion allows fluid PL to regain the surface through the fusion pores. This mechanism explains both the rapid reincorporation of surfactant PL into the monolayer during adsorption and during film expansion and the progressive improvement in surface activity during repeated compression.

The interfacial surface tension of the lung is regulated by phospholipid-rich pulmonary surfactant films. Small changes in temperature affect surfactant structure and function in vitro. We compared the compositional, thermodynamic and functional properties of surfactant from hibernating and summer-active 13-lined ground squirrels (Ictidomys tridecemlineatus) with porcine surfactant to understand structure-function relationships in surfactant membranes and films. Hibernating squirrels had more surfactant large aggregates with more fluid monounsaturated molecular species than summer-active animals. The latter had more unsaturated species than porcine surfactant. Cold-adapted surfactant membranes displayed gel-to-fluid transitions at lower phase transition temperatures with reduced enthalpy. Both hibernating and summer-active squirrel surfactants exhibited lower enthalpy than porcine surfactant. LAURDAN fluorescence and DPH anisotropy revealed that surfactant bilayers from both groups of squirrels possessed similar ordered phase characteristics at low temperatures. While ground squirrel surfactants functioned well during dynamic cycling at 3, 25, and 37 °C, porcine surfactant demonstrated poorer activity at 3 °C but was superior at 37 °C. Consequently the surfactant composition of ground squirrels confers a greater thermal flexibility relative to homeothermic mammals, while retaining tight lipid packing at low body temperatures. This may represent the most critical feature contributing to sustained stability of the respiratory interface at low lung volumes. Thus, while less effective than porcine surfactant at 37 °C, summer-active surfactant functions adequately at both 37 °C and 3 °C allowing these animals to enter hibernation. Here further compositional alterations occur which improve function at low temperatures by maintaining adequate stability at low lung volumes and when temperature increases during arousal from hibernation.

Background The lung and thyroid are derived from the anterior endoderm. Retinoic acid and Fgf signalling are known to be essential for development of the lung in mouse but little is known on how the lung and thyroid are specified in Xenopus . Results If either retinoic acid or Fgf signalling is inhibited, there is no differentiation of the lung as assayed by expression of sftpb . There is no change in expression of thyroid gland markers when retinoic acid signalling is blocked after gastrulation and when Fgf signalling is inhibited there is a short window of time where pax2 expression is inhibited but expression of other markers is unaffected. If exogenous retinoic acid is given to the embryo between embryonic stages 20 and 26, the presumptive thyroid expresses sftpb and sftpc , specific markers of lung differentiation and expression of key thyroid transcription factors is lost. When the presumptive thyroid is transplanted into the posterior embryo, it also expresses sftpb , although pax2 expression is not blocked. Conclusions After gastrulation, retinoic acid is required for lung but not thyroid differentiation in Xenopus while Fgf signalling is needed for lung but only for early expression of pax2 in the thyroid. Exposure to retinoic acid can cause the presumptive thyroid to switch to a lung developmental program.

Pulmonary surfactant, a mixture of lipids and proteins, regulates the surface tension at the air-liquid interface of the lung. Reduced body temperature during hibernation is accompanied in 13-lined ground squirrels (Ictidomys tridecemlineatus) by an increase in fluid monounsaturated phosphatidylcholine (PC) species (e.g. PC 16:0/16:1, PC 16:0/18:1) and phosphatidylglycerol (PG) species (e.g. PG 16:0/18:1, PG 18:0/18:2), but fewer disaturated PC and PG species. Previously we speculated that altered surfactant lipid composition during metabolic depression states such as torpor or hibernation will reduce the phase transition temperature (Tm) of the mixture, enabling pulmonary surfactant to remain fluid over a broader range of temperatures and thereby maintaining respiratory function. Here we analyze thermodynamic properties and behavior of surfactant from hibernating and summer-active 13-lined ground squirrels in relation to natural porcine surfactant, using differential scanning calorimetry and LAURDAN fluorescence spectroscopy. In addition we conducted epifluorescence studies to visualize changes in phase coexistence of surfactant films of hibernating and summer-active animals. Surfactant membranes of hibernators showed gel-to-fluid transitions at lower Tm with reduced enthalpy relative to membranes from summer-active squirrels. Both exhibited lower enthalpy than porcine surfactant. LAURDAN fluorescence and epifluoresence suggested possible structural re-arrangements of surfactant membrane lipids and films, respectively, in hibernators. These exhibited a similarly dehydrated and condensed highly packed ordered phase as for summer active squirrels, despite differences in composition and Tm. In conclusion, pulmonary surfactant composition changes in hibernating squirrels to increase overall fluidity, but to maintain an ordered membrane structure at low temperature.

Pulmonary surfactant is a complex lipid-protein mixture whose main function is to reduce the surface tension at the air-liquid interface of alveoli to minimize the work of breathing. The exact mechanism by which surfactant monolayers and multilayers are formed and how they lower surface tension to very low values during lateral compression remains uncertain. We used time-of-flight secondary ion mass spectrometry to study the lateral organization of lipids and peptide in surfactant preparations ranging in complexity. We show that we can successfully determine the location of phospholipids, cholesterol and a peptide in surfactant Langmuir-Blodgett films and we can determine the effect of cholesterol and peptide addition. A thorough understanding of the lateral organization of PS interfacial films will aid in our understanding of the role of each component as well as different lipid-lipid and lipid-protein interactions. This may further our understanding of pulmonary surfactant function.

Pulmonary surfactant facilitates breathing by forming a surface tension reducing film at the air-liquid interface of the alveoli. The objective was to characterize the structure of surfactant films using endogenous rat surfactant. Solid-support surfactant films, at different surface pressures, were obtained using a Langmuir balance and were analyzed using atomic force microscopy. The results showed a lipid film structure with three distinct phases: liquid expanded, liquid ordered and liquid condensed. The area covered by the liquid condensed domains increased as surface pressure increased. The presence of liquid ordered phase within these structures correlated with the cholesterol content. At a surface pressure of 50 mN/m, stacks of bilayers appeared. Several structural details of these films differ from previous observations made with goat and exogenous surfactants. Overall, the data indicate that surfactant films demonstrate phase separation at low surface pressures and multilayer formation at higher pressure, features likely important for normal surfactant function.

The ability of pulmonary surfactant to drive surface tension (ST) to low values near 0 mN/m during compression stabilizes the alveoli at end expiration. The manner in which surfactant phospholipids (PL) (50% unsaturation) attain surface tension near zero is still not clear. Atomic force microscopy (AFM) of rat surfactant extract films at a surface pressure (SP) of 20 mN/m reveals microdomains (mD), apparently composed of liquid-ordered (L0) and tilted-condensed (TC) phases within the predominant liquid-expanded (LE) phase. Numerous nanodomains (nD) were also present. As SP increased to 40 mN/m, the area composed of mD increased but the nD decreased. This contrasts with previous studies with bovine surfactant where the mD appeared to form nD. At 50 mN/m, the film exhibited the formation of numerous stacked multilayers (PL bilayers) which could be incorporated into the monolayer during film expansion. The present results are consistent with the reversible squeeze-out of unsaturated PL into multilayers during compression, resulting in a monolayer highly enriched in gel phase saturated PL components. Furthermore, as with other surfactants, the presence of L0 phase appears related to cholesterol. Reducing the cholesterol content by acetone precipitation resulted in the loss of L0 phase, further implicating this sterol in surfactant phase separation.

Recent studies have demonstrated intraventricular infusions of propionic acid (PPA) a dietary and enteric short-chain fatty acid can produce brain and behavioral changes similar to those observed in autism spectrum disorder (ASD). The effects of PPA were further evaluated to determine if there are any alterations in brain lipids associated with the ASD-like behavioral changes observed following intermittent intraventricular infusions of PPA, the related enteric metabolite butyric acid (BUT) or phosphate-buffered saline vehicle. Both PPA and BUT produced significant increases (p < 0.001) in locomotor activity (total distance travelled and stereotypy). PPA and to a lesser extent BUT infusions decreased the levels of total monounsaturates, total omega 6 fatty acids, total phosphatidylethanolamine plasmalogens, the ratio of omega 6 : omega 3 and elevated the levels of total saturates in separated phospholipid species. In addition, total acylcarnitines, total long-chain (C12-C24) acylcarnitines, total short-chain (C2 to C9) acylcarnitines, and the ratio of bound to free carnitine were increased following infusions with PPA and BUT. These results provide evidence of a relationship between changes in brain lipid profiles and the occurrence of ASD-like behaviors using the autism rodent model. We propose that altered brain fatty acid metabolism may contribute to ASD.