Molecular confirmation of nine cases of Cornelia de Lange syndrome diagnosed prenatally.

OBJECTIVES: Cornelia de Lange syndrome (CdLS) is characterized by distinct facial features, growth retardation, upper limb reduction defects, hirsutism, and intellectual disability. NIPBL mutations have been identified in approximately 60% of patients with CdLS diagnosed postnatally. Prenatal ultrasound findings include upper limb reduction defects, intrauterine growth restriction, and micrognathia. CdLS has also been associated with decreased PAPP-A and increased nuchal translucency (NT). We reviewed NIPBL sequence analysis results for 12 prenatal samples in our laboratory to determine the frequency of mutations in our cohort. METHODS: This retrospective study analyzed data from all 12 prenatal cases with suspected CdLS, which were received by The University of Chicago Genetic Services Laboratories. Diagnostic NIPBL sequencing was performed for all samples. Clinical information was collected from referring physicians. RESULTS: NIPBL mutations were identified in 9 out of the 12 cases prenatally (75%). Amongst the NIPBL mutation-positive cases with clinical information available, the most common findings were upper limb malformations and micrognathia. Five patients had NT measurements in the first trimester, of which four were noted to be increased. CONCLUSION: We demonstrate that prenatally-detected phenotypes of CdLS, particularly severe micrognathia and bilateral upper limb defects, are associated with an increased frequency of NIPBL mutations.

Comparison of pharmacodynamics between insulin glargine 100 U/mL and insulin glargine 300 U/mL in healthy cats.

Insulin glargine (IGla) is a synthetic human-recombinant insulin analog that is used routinely in people as a q24h basal insulin. The 300 U/mL (U300) formulation of IGla is associated with longer duration of action and less within-day variability, making it a better basal insulin compared with the 100 U/mL (U100) formulation. We hypothesized that in healthy cats, IGlaU300 has a flatter time-action profile and longer duration of action compared with IGlaU100. Seven healthy neutered male, purpose-bred cats were studied in a randomized, crossover design. Pharmacodynamics of IGlaU100 and IGlaU300 (0.8 U/kg, subcutaneous) were determined by the isoglycemic clamp method. The time-action profile of IGlaU300 was flatter compared with IGlaU100 as demonstrated by lower peak (5.6 ± 1.1 mg/kg/min vs 8.3 ± 1.9 mg/kg/min, respectively; P = 0.04) with no difference in total metabolic effect (ME; P = 0.7) or duration of action (16.8 h ± 4.7 h vs 13.4 h ± 2.6 h; P = 0.2). The greater fraction of ME in the 12- to 24-h period postinjection (35 ± 23% vs 7 ± 8% respectively; P = 0.048) and lower intraday GIR% variability (7.8 ± 3.7% vs 17.4 ± 8.2% respectively; P = 0.03) supports a flatter time-action profile of IGlaU300. There were no differences in onset and end of the action. In summary, although both formulations have a similar duration of action that is well below 24 h, the ME of IGlaU300 is more evenly distributed over a 24 h period in healthy cats, making it a better candidate for once-daily injection in diabetics compared with IGlaU100.

The Mars 2020 Engineering Cameras and Microphone on the Perseverance Rover: A Next-Generation Imaging System for Mars Exploration.

The Mars 2020 Perseverance rover is equipped with a next-generation engineering camera imaging system that represents an upgrade over previous Mars rover missions. These upgrades will improve the operational capabilities of the rover with an emphasis on drive planning, robotic arm operation, instrument operations, sample caching activities, and documentation of key events during entry, descent, and landing (EDL). There are a total of 16 cameras in the Perseverance engineering imaging system, including 9 cameras for surface operations and 7 cameras for EDL documentation. There are 3 types of cameras designed for surface operations: Navigation cameras (Navcams, quantity 2), Hazard Avoidance Cameras (Hazcams, quantity 6), and Cachecam (quantity 1). The Navcams will acquire color stereo images of the surface with a 96 ∘ × 73 ∘ field of view at 0.33 mrad/pixel. The Hazcams will acquire color stereo images of the surface with a 136 ∘ × 102 ∘ at 0.46 mrad/pixel. The Cachecam, a new camera type, will acquire images of Martian material inside the sample tubes during caching operations at a spatial scale of 12.5 microns/pixel. There are 5 types of EDL documentation cameras: The Parachute Uplook Cameras (PUCs, quantity 3), the Descent stage Downlook Camera (DDC, quantity 1), the Rover Uplook Camera (RUC, quantity 1), the Rover Descent Camera (RDC, quantity 1), and the Lander Vision System (LVS) Camera (LCAM, quantity 1). The PUCs are mounted on the parachute support structure and will acquire video of the parachute deployment event as part of a system to characterize parachute performance. The DDC is attached to the descent stage and pointed downward, it will characterize vehicle dynamics by capturing video of the rover as it descends from the skycrane. The rover-mounted RUC, attached to the rover and looking upward, will capture similar video of the skycrane from the vantage point of the rover and will also acquire video of the descent stage flyaway event. The RDC, attached to the rover and looking downward, will document plume dynamics by imaging the Martian surface before, during, and after rover touchdown. The LCAM, mounted to the bottom of the rover chassis and pointed downward, will acquire 90 ∘ × 90 ∘ FOV images during the parachute descent phase of EDL as input to an onboard map localization by the Lander Vision System (LVS). The rover also carries a microphone, mounted externally on the rover chassis, to capture acoustic signatures during and after EDL. The Perseverance rover launched from Earth on July 30th, 2020, and touchdown on Mars is scheduled for February 18th, 2021.

Protein translocation at the ER membrane: A complex process becomes more so.

Protein transport across or insertion into a membrane is facilitated by multicomponent protein complexes that reside in the bilayer. Current models propose that these complexes mediate translocation and integration by an obligate sequence of interactions between the substrate polypeptide and other components. Recent discoveries extend and complicate these models, but, more importantly, they remind us that our current level of understanding of the actual molecular mechanisms involved is crude and represents only the tip of the iceberg.

Protein translocation across the endoplasmic reticulum membrane: identification by photocross-linking of a 39-kD integral membrane glycoprotein as part of a putative translocation tunnel.

The molecular environment of secretory proteins during translocation across the ER membrane was examined by photocross-linking. Nascent preprolactin chains of various lengths, synthesized by in vitro translation of truncated messenger RNAs in the presence of N epsilon-(5-azido-2-nitrobenzoyl)-Lys-tRNA, signal recognition particle, and microsomal membranes, were used to position photoreactive probes at various locations within the membrane. Upon photolysis, each nascent chain species was cross-linked to an integral membrane glycoprotein with a deduced mass of 39 kD (mp39) via photoreactive lysines located in either the signal sequence or the mature prolactin sequence. Thus, different portions of the nascent preprolactin chain are in close proximity to the same membrane protein during the course of translocation, and mp39 therefore appears to be part of the translocon, the specific site of protein translocation across the ER membrane. The similarity of the molecular and cross-linking properties of mp39 and the glyco-protein previously identified as a signal sequence receptor (Wiedmann, M., T. V. Kurzchalia, E. Hartmann, and T. A. Rapoport. 1987. Nature [Lond.]. 328: 830-833) suggests that these two proteins may be identical. Our data indicate, however, that mp39 does not (or not only) function as a signal sequence receptor, but rather may be part of a putative translocation tunnel.

The interaction of the chaperonin tailless complex polypeptide 1 (TCP1) ring complex (TRiC) with ribosome-bound nascent chains examined using photo-cross-linking.

The eukaryotic chaperonin tailless complex polypeptide 1 (TCP1) ring complex (TRiC) (also called chaperonin containing TCP1 [CCT]) is a hetero-oligomeric complex that facilitates the proper folding of many cellular proteins. To better understand the manner in which TRiC interacts with newly translated polypeptides, we examined its association with nascent chains using a photo-cross-linking approach. To this end, a series of ribosome-bound nascent chains of defined lengths was prepared using truncated mRNAs. Photoactivatable probes were incorporated into these (35)S- labeled nascent chains during translation. Upon photolysis, TRiC was cross-linked to ribosome-bound polypeptides exposing at least 50-90 amino acids outside the ribosomal exit channel, indicating that the chaperonin associates with much shorter nascent chains than indicated by previous studies. Cross-links were observed for nascent chains of the cytosolic proteins actin, luciferase, and enolase, but not to ribosome-bound preprolactin. The pattern of cross-links became more complex as the nascent chain increased in length. These results suggest a chain length-dependent increase in the number of TRiC subunits involved in the interaction that is consistent with the idea that the substrate participates in subunit-specific contacts with the chaperonin. Both ribosome isolation by centrifugation through sucrose cushions and immunoprecipitation with anti-puromycin antibodies demonstrated that the photoadducts form on ribosome-bound polypeptides. Our results indicate that TRiC/CCT associates with the translating polypeptide shortly after it emerges from the ribosome and suggest a close association between the chaperonin and the translational apparatus.

Two intermembrane space TIM complexes interact with different domains of Tim23p during its import into mitochondria.

Tim23p (translocase of the inner membrane) is an essential import component located in the mitochondrial inner membrane. To determine how the Tim23 protein itself is transported into mitochondria, we used chemical cross-linking to identify proteins adjacent to Tim23p during its biogenesis. In the absence of an inner membrane potential, Tim23p is translocated across the mitochondrial outer membrane, but not inserted into the inner membrane. At this intermediate stage, we find that Tim23p forms cross-linked products with two distinct protein complexes of the intermembrane space, Tim8p-Tim13p and Tim9p-Tim10p. Tim9p and Tim10p cross-link to the COOH-terminal domain of the Tim23 protein, which carries all of the targeting signals for Tim23p. Therefore, our results suggest that the Tim9p-Tim10p complex plays a key role in Tim23p import. In contrast, Tim8p and Tim13p cross-link to the hydrophilic NH(2)-terminal segment of Tim23p, which does not carry essential import information and, thus, the role of Tim8p-Tim13p is unclear. Tim23p contains two matrix-facing, positively charged loops that are essential for its insertion into the inner membrane. The positive charges are not required for interaction with the Tim9p-Tim10p complex, but are essential for cross-linking of Tim23p to components of the inner membrane insertion machinery, including Tim54p, Tim22p, and Tim12p.

A nascent membrane protein is located adjacent to ER membrane proteins throughout its integration and translation.

The immediate environment of nascent membrane proteins undergoing integration into the ER membrane was investigated by photocrosslinking. Nascent polypeptides of different lengths, each containing a single IgM transmembrane sequence that functions either as a stop-transfer or a signal-anchor sequence, were synthesized by in vitro translation of truncated mRNAs in the presence of N epsilon-(5-azido-2-nitrobenzoyl)-Lys-tRNA, signal recognition particle, and microsomal membranes. This yielded nascent chains with photoreactive probes at one end of the transmembrane sequence where two lysine residues are located. When irradiated, these nascent chains reacted covalently with several ER proteins. One prominent crosslinking target was a glycoprotein similar in size to a protein termed mp39, shown previously to be situated adjacent to a secretory protein during its translocation across the ER membrane (Krieg, U. C., A. E. Johnson, and P. Walter. 1989. J. Cell Biol. 109:2033-2043; Wiedmann, M., D. Goerlich, E. Hartmann, T. V. Kurzchalia, and T. A. Rapoport. 1989. FEBS (Fed. Eur. Biochem. Soc.) Lett. 257:263-268) and likely to be identical to a protein previously designated the signal sequence receptor (Wiedmann, M., T. V. Kurzchalia, E. Hartmann, and T. A. Rapoport. 1987. Nature (Lond.). 328:830-833). Changing the orientation of the transmembrane domain in the bilayer, or making the transmembrane domain the first topogenic sequence in the nascent chain instead of the second, did not significantly alter the identities of the ER proteins that were the primary crosslinking targets. Furthermore, the nascent chains crosslinked to the mp39-like glycoprotein and other microsomal proteins even after the cytoplasmic tail of the nascent chain had been lengthened by nearly 100 amino acids beyond the stop-transfer sequence. Yet when the nascent chain was allowed to terminate normally, the major photocrosslinks were no longer observed, including in particular that to the mp39-like glycoprotein. These results show that the transmembrane segment of a nascent membrane protein is located adjacent to the mp39-like glycoprotein and other ER proteins during the integration process, and that at least a portion of the nascent chain remains in close proximity to these ER proteins until translation has been completed.

Role of 4.5S RNA in assembly of the bacterial signal recognition particle with its receptor.

The mechanism by which a signal recognition particle (SRP) and its receptor mediate protein targeting to the endoplasmic reticulum or to the bacterial plasma membrane is evolutionarily conserved. In Escherichia coli, this reaction is mediated by the Ffh/4.5S RNA ribonucleoprotein complex (Ffh/4.5S RNP; the SRP) and the FtsY protein (the SRP receptor). We have quantified the effects of 4.5S RNA on Ffh-FtsY complex formation by monitoring changes in tryptophan fluorescence. Surprisingly, 4.5S RNA facilitates both assembly and disassembly of the Ffh-FtsY complex to a similar extent. These results provide an example of an RNA molecule facilitating protein-protein interactions in a catalytic fashion.

The translocon: more than a hole in the ER membrane?

In eukaryotes, the vast majority of secreted and integral membrane proteins are targeted to the membrane of the endoplasmic reticulum (ER) early during translation. These polypeptides are then either transported across or inserted into the ER membrane at sites termed translocons. As protein translocation occurs through an aqueous pore, the minimal requirement for a translocon is a passive structure that provides a passage-way across the membrane. However, recent data suggest that the translocon is a complex structure that orchestrates the localization, orientation, maturation and possibly degradation of nascent chains.

Protein translocation: is Hsp70 pulling my chain?

Hsp70 proteins in the lumen of the endoplasmic reticulum and in the mitochondrial matrix are thought to drive the translocation of proteins into each organelle. Recent experiments aimed at distinguishing between two models for Hsp70 function appear to reach opposite conclusions.

AF Classification from a Short Single Lead ECG Recording: the PhysioNet/Computing in Cardiology Challenge 2017.

The PhysioNet/Computing in Cardiology (CinC) Challenge 2017 focused on differentiating AF from noise, normal or other rhythms in short term (from 9-61 s) ECG recordings performed by patients. A total of 12,186 ECGs were used: 8,528 in the public training set and 3,658 in the private hidden test set. Due to the high degree of inter-expert disagreement between a significant fraction of the expert labels we implemented a mid-competition bootstrap approach to expert relabeling of the data, levering the best performing Challenge entrants' algorithms to identify contentious labels. A total of 75 independent teams entered the Challenge using a variety of traditional and novel methods, ranging from random forests to a deep learning approach applied to the raw data in the spectral domain. Four teams won the Challenge with an equal high F1 score (averaged across all classes) of 0.83, although the top 11 algorithms scored within 2% of this. A combination of 45 algorithms identified using LASSO achieved an F1 of 0.87, indicating that a voting approach can boost performance.

Toxicologic aspects of photosensitization in livestock.

Two types of plant-caused photosensitizations are recognized in livestock: 1) primary, wherein the phototoxic agent in the plant is ingested and reaches the skin chemically unchanged; and 2) secondary, wherein the phototoxic agent in the porphyrin phylloerythrin produced by chlorophyll degradation in ruminant stomachs. Phylloerythrin is normally excreted in bile but is allowed to reach the skin when hepatic damage interferes with the phylloerythrin-excreting mechanism. Primary photosensitizing plant toxins are few, whereas secondary photosensitizations can be caused by damage to the liver by a variety of plant and other toxins. Plants causing each type of photosensitization are discussed and clinical manifestations of the disease in livestock are summarized. Tetradymia species are one of the most economically important causes of phototoxicity in livestock. The etiology of this phototoxic syndrome in sheep and the importance of sagebrush species as preconditioning agents for phototoxicity are discussed.

The cholesterol-dependent cytolysins.

In view of the recent studies on the CDCs, a reasonable schematic of the stages leading to membrane insertion of the CDCs can be assembled. As shown in Fig. 3, we propose that the CDC first binds to the membrane as a monomer. These monomers then diffuse laterally on the membrane surface to encounter other monomers or incomplete oligomeric complexes. Presumably, once the requisite oligomer size is reached, the prepore complex is converted into the pore complex and a large membrane channel is formed. During the conversion of the prepore complex to the pore complex, we predict that the TMHs of the subunits in the prepore complex insert into the bilayer in a concerted fashion to form the large transmembrane beta-barrel, although this still remains to be confirmed experimentally. Many intriguing problems concerning the cytolytic mechanism of the CDCs remain unsolved. The nature of the initial interaction of the CDC monomer with the membrane is currently one of the most controversial questions concerning the CDC mechanism. Is cholesterol involved in this interaction, as previously assumed, or do specific receptors exist for these toxins that remain to be discovered? Also, the trigger for membrane insertion and the regions of these toxins that facilitate the [figure: see text] interaction of the monomers during prepore complex formation are unknown. In addition, the temporal sequence of the multiple structural changes that accompany the conversion of the soluble CDC monomer into a membrane-inserted oligomer have yet to be defined or characterized kinetically.

Effects of propranolol on phosphatidate phosphohydrolase and mitogen-activated protein kinase activities in A7r5 vascular smooth muscle cells.

High doses of propranolol inhibit phosphatidate phosphohydrolase (PAP) activity in intact cells, thus blocking metabolism of phosphatidic acid (PA), product of the phospholipase D (PLD) reaction. Vasopressin and phorbol ester activate PLD and ERK (extracellular signal-regulated protein kinase) mitogen-activated protein kinases in A7r5, a rat vascular smooth muscle cell line. Propranolol increased PA levels in intact A7r5 cells and inhibited cytosolic PAP and membrane calcium-independent phospholipase A2 but did not activate PLD or enhance agonist-induced PA accumulation. Incubation of cells with 200 microM propranolol for 10-45 min markedly elevated PA but caused only partial activation of ERKs. Propranolol and other lipophilic amines caused a time- and dose-dependent detachment of cells from their substrate. These results confirm that elevation of PA is not a strong signal for ERK activation and emphasize that caution should be exercised in using propranolol as a PAP inhibitor in intact cells.

Anatomical localization of the effects of 17 beta-estradiol on oxytocin receptor binding in the ventromedial hypothalamic nucleus.

Oxytocin (OT) neurotransmission plays a role in the facilitation of steroid-dependent sexual receptivity in the rat. One way in which the ovarian steroid 17 beta-estradiol (E2) has been shown to modulate OT transmission is by increasing OT receptor binding in certain brain areas involved in the regulation of female sexual behavior such as the ventromedial hypothalamic nucleus (VMN). This study was undertaken to describe the distribution of OT receptors within the VMN that are regulated by physiological levels of E2. With quantitative autoradiographic methods, we measured [3H]OT binding in ovariectomized female rats implanted with Silastic capsules containing cholesterol, 5% E2, or 100% E2. In addition, plasma E2 levels, pituitary progestin receptor binding, and uterine weights were measured in animals from each treatment group. Results of this study showed that physiological levels of E2 increased [3H]OT binding in caudal regions of the ventrolateral VMN and stimulated maximal uterine growth and pituitary progestin receptor binding. However, in more rostral VMN sections, E2 induced a dose-dependent increase in [3H]OT binding. These data suggest that ovarian steroids sensitize the brain to OT by increasing OT receptor binding in certain brain areas involved in the regulation of sexual receptivity.

The regulation of oxytocin receptor binding in the ventromedial hypothalamic nucleus by testosterone and its metabolites.

Oxytocin (OT) receptor binding in the ventromedial hypothalamic nucleus is regulated by testosterone (T) in male rats. However, T is metabolized in the brain, and many of the central effects of T are mediated by its metabolites. The experiments reported here were designed to determine whether T affects OT receptor binding directly or through the action of its metabolites 17 beta-estradiol and 5 alpha-dihydrotestosterone. Adult male rats were either sham operated or castrated and treated 1 week later with T propionate (TP), 17 beta-estradiol benzoate (EB), dihydrotestosterone benzoate (DHTB), DHTB plus EB, or oil. OT receptor binding was assessed autoradiographically using [125I]d(CH2)5[Tyr(Me)2,Thr4,Tyr-NH2(9)]OVT. In addition, seminal vesicle weights were measured as an index of androgenic activity. These experiments showed that TP and DHTB plus EB increased OT receptor binding in the ventromedial hypothalamic nucleus to the levels in intact males. Treatment with EB alone partially reinstated binding to the levels in intact males, while DHTB treatment was without effect. Castrated males treated with either TP or DHTB had seminal vesicle weights comparable to those of gonadally intact males and greater than those of animals in all other steroid conditions, indicating that sufficient levels of circulating steroids were attained in these groups. These data suggest that the induction of hypothalamic OT receptor binding by T is the result of the combined actions of estradiol and dihydrotestosterone. However, the mechanism underlying this interaction is unknown.

Time course of the estradiol-dependent induction of oxytocin receptor binding in the ventromedial hypothalamic nucleus of the rat.

Oxytocin (OT) transmission is involved in the steroid-dependent display of sexual receptivity in rats. One of the biochemical processes stimulated by the ovarian steroid 17 beta-estradiol (E2) that is relevant to reproduction is the induction of OT receptor binding in the ventromedial hypothalamic nucleus (VMN). The purpose of these experiments was to determine if E2-induced changes in OT receptor binding in the VMN occur within a time frame relevant to cyclic changes in ovarian steroid secretion. OT receptor binding was measured in the VMN of ovariectomized rats implanted for 0-96 h with E2-containing Silastic capsules. The rate of decay of OT receptor binding was measured in another group of animals 6-48 h after capsule removal. Receptors were labeled with the specific OT receptor antagonist [125I]d(CH2)5[Tyr(Me)2,Thr4,Tyr-NH2(9)]OVT, and binding was measured with quantitative autoradiographic methods. In addition, plasma E2 levels and uterine weights were assessed in animals from each treatment condition. Significant increases in E2-dependent OT receptor binding and uterine weight occurred within 24 h of steroid treatment. After E2 withdrawal, OT receptor binding and uterine weight decreased significantly within 24 h. These results are consistent with the hypothesis that steroid modulation of OT receptor binding is necessary for the induction of sexual receptivity.

Cholecystokinin, dopamine D2 and N-methyl-D-aspartate binding sites in the nucleus of the solitary tract of the rat: possible relationship to ingestive behavior.

Receptor autoradiography was used to investigate the distribution of brainstem binding sites for cholecystokinin, dopamine and N-methyl-D-aspartate with particular reference to the nucleus of the solitary tract of the rat, an area involved in the control of ingestive behavior. Binding sites for the A and B subtypes of the cholecystokinin receptor, labeled with [(125)I]cholecystokinin octapeptide sulfate in the presence or absence of antagonists for the devazepide (A) or L-365,260 (B) receptor, were present throughout the caudal rostral extent of the nucleus of the solitary tract, the A type predominating in the commissural, medial and gelatinous part and the B type in the lateral part. In the most rostral part of the medial nucleus of the solitary tract, both A and B receptors were present. Dopamine D2 receptors, labeled with [(125)I]NCQ-298, were found in all parts of the nucleus of the solitary tract. No binding to the dopamine D1 receptor, labeled with [(125)I]SCH-23982, was found in the brainstem. N-Methyl-D-aspartate receptors, labeled with [(3)H]dizocilpine maleate, were also present in the entire caudorostral extent of the nucleus of the solitary tract. Binding to cholecystokinin A receptors was co-distributed with [(125)I]NCQ-298 and [(3)H]dizocilpine maleate binding in the caudal and rostral parts of the nucleus of the solitary tract, and binding to cholecystokinin B receptors overlapped with [(125)I]NCQ-298 and [(3)H]dizocilpine maleate binding in the rostral nucleus of the solitary tract. These results are consistent with the hypothesis that cholecystokinin, dopamine and glutamate interact in the nucleus of the solitary tract in the control of ingestive behavior.

Chronic treatment with a classical neuroleptic alters excitatory amino acid and GABAergic neurotransmission in specific regions of the rat brain.

The purpose of the following experiments was to describe some of the neurochemical changes that occur in the basal ganglia of rats exposed chronically to a classical neuroleptic, fluphenazine, and to relate these changes to extrapyramidal motor dysfunction. For these studies a combination of behavioural, receptor autoradiographic and in situ hybridization methods were employed. Preliminary pharmacological studies on GABA receptors showed that incubation in Tris-acetate rather than Tris-citrate buffer increased the number of binding sites labelled by [3H]muscimol by over 120% without affecting binding affinity or selectivity. The results of experiments with fluphenazine showed that treatment for six months increased the frequency of vacuous chewing movements compared to controls. In the striatum, changes in GABA transmission were observed in fluphenazine-treated rats with increases in glutamate decarboxylase mRNA levels in the caudate nucleus, dorsal shell and core of the accumbens and decreases in [3H]muscimol binding in the caudate and dorsal shell regions. These data suggest that fluphenazine treatment increased GABA transmission in specific subregions of the caudate and accumbens nuclei. In addition, glutamate decarboxylase mRNA levels were elevated in the entopeduncular nucleus of fluphenazine-treated animals. Autoradiographic analysis of excitatory amino acid binding showed that fluphenazine exposure decreased [3H]alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid binding in entopeduncular nucleus and in the ventrolateral thalamic nucleus and decreased [3H]dizocilpine maleate binding in the medial geniculate nucleus. These experiments show that in addition to altering GABA transmission, chronic neuroleptic exposure alters excitatory amino acid transmission in specific regions of the basal ganglia-thalamocortical motor system. The neuroleptic dependent increases in glutamate decarboxylase mRNA levels in the entopeduncular nucleus may reflect changes in neurotransmission in the indirect pathway connecting the major input and output nuclei of the basal ganglia. Changes in some of these brain regions may be related to the occurrence of extrapyramidal motor disturbances.