Using poly(vinyldodecylimidazolium bromide) for the in-situ product recovery of n-butanol.

The mitigation of end-product inhibition during the biosynthesis of n-butanol is demonstrated for an in-situ product recovery (ISPR) system employing a poly(ionic liquid) (PIL) absorbent. The thermodynamic affinity of poly(vinyldodecylimidazolium bromide) [P(VC12 ImBr)] for n-butanol, acetone and ethanol versus water was measured at conditions experienced in a typical acetone-ethanol-butanol (ABE) fermentation. In addition to providing a high n-butanol partition coefficient (PC = 6.5) and selectivity (αBuOH/water = 46), P(VC12 ImBr) is shown to be biocompatible with Saccharomyces cerevisiae and Clostridium acetobutylicum. Furthermore, the diffusivity of n-butanol in a hydrated PIL provides absorption rates that support ISPR applications. Using a 5 wt% PIL phase fraction relative to the aqueous phase mass, P(VC12 ImBr) improved the volumetric productivity of a batch ABE ISPR process by 31% relative to a control fermentation. The concentration of n-butanol in the P(VC12 ImBr) phase was sufficient to increase the alcohol concentration from 1.5 wt% in the fermentation medium to 25 wt% in the saturated PIL, thereby facilitating downstream n-butanol recovery.

Axonal plasticity of age-defined dentate granule cells in a rat model of mesial temporal lobe epilepsy.

Dentate granule cell (DGC) mossy fiber sprouting (MFS) in mesial temporal lobe epilepsy (mTLE) is thought to underlie the creation of aberrant circuitry which promotes the generation or spread of spontaneous seizure activity. Understanding the extent to which populations of DGCs participate in this circuitry could help determine how it develops and potentially identify therapeutic targets for regulating aberrant network activity. In this study, we investigated how DGC birthdate influences participation in MFS and other aspects of axonal plasticity using the rat pilocarpine-induced status epilepticus (SE) model of mTLE. We injected a retrovirus (RV) carrying a synaptophysin-yellow fluorescent protein (syp-YFP) fusion construct to birthdate DGCs and brightly label their axon terminals, and compared DGCs born during the neonatal period with those generated in adulthood. We found that both neonatal and adult-born DGC populations participate, to a similar extent, in SE-induced MFS within the dentate gyrus inner molecular layer (IML). SE did not alter hilar MF bouton density compared to sham-treated controls, but adult-born DGC bouton density was greater in the IML than in the hilus after SE. Interestingly, we also observed MF axonal reorganization in area CA2 in epileptic rats, and these changes arose from DGCs generated both neonatally and in adulthood. These data indicate that both neonatal and adult-generated DGCs contribute to axonal reorganization in the rat pilocarpine mTLE model, and indicate a more complex relationship between DGC age and participation in seizure-related plasticity than was previously thought.

Selecting polymers for two-phase partitioning bioreactors (TPPBs): Consideration of thermodynamic affinity, crystallinity, and glass transition temperature.

Two-phase partitioning bioreactor technology involves the use of a secondary immiscible phase to lower the concentration of cytotoxic solutes in the fermentation broth to subinhibitory levels. Although polymeric absorbents have attracted recent interest due to their low cost and biocompatibility, material selection requires the consideration of properties beyond those of small molecule absorbents (i.e., immiscible organic solvents). These include a polymer's (1) thermodynamic affinity for the target compound, (2) degree of crystallinity (wc ), and (3) glass transition temperature (Tg ). We have examined the capability of three thermodynamic models to predict the partition coefficient (PC) for n-butyric acid, a fermentation product, in 15 polymers. Whereas PC predictions for amorphous materials had an average absolute deviation (AAD) of ≥16%, predictions for semicrystalline polymers were less accurate (AAD ≥ 30%). Prediction errors were associated with uncertainties in determining the degree of crystallinity within a polymer and the effect of absorbed water on n-butyric acid partitioning. Further complications were found to arise for semicrystalline polymers, wherein strongly interacting solutes increased the polymer's absorptive capacity by actually dissolving the crystalline fraction. Finally, we determined that diffusion limitations may occur for polymers operating near their Tg , and that the Tg can be reduced by plasticization by water and/or solute. This study has demonstrated the impact of basic material properties that affects the performance of polymers as sequestering phases in TPPBs, and reflects the additional complexity of polymers that must be taken into account in material selection.

Brainstem auditory evoked responses in foals: reference values, effect of age, rate of acoustic stimulation, and neurologic deficits.

BACKGROUND: Age and rate of acoustic stimulation affect peak latencies in brainstem auditory evoked responses (BAER) in humans. Those effects are unknown in foals. HYPOTHESIS/OBJECTIVES: Our goals were to (1) establish reference values for BAER in foals by using 3 different stimulation protocols, (2) evaluate the effects of age and stimulation frequencies on BAER tracing in foals up to 6 months old, and (3) compare the data with BAER obtained from foals with central nervous system (CNS) disorders. ANIMALS: Thirty-nine neurologically normal foals and 16 foals with neurologic diseases. METHODS: Prospective observational clinical study. BAER recorded by using 3 protocols of stimulation (11.33 repetitions per second [Hz]/70 decibel normal hearing level [dBNHL]; 11.33 Hz/90 dBNHL; 90 Hz/70 dBNHL). RESULTS: No effect of age was observed in normal foals (P > .005). No significant difference was observed for latencies and interpeak latencies (IPL) when comparing foals with neurologic diseases and normal foals (P > .05), but 78.6% of foals with neurologic diseases had an asymmetry in their tracing, reflecting a difference in conduction time between the left and right side of the brainstem. Increasing the stimulation rate did not improve detection of CNS disorders. CONCLUSIONS AND CLINICAL IMPORTANCE: We propose BAER reference values for foals up to 6 months of age by using 3 protocols. Most foals with neurologic deficits had abnormal BAER tracing.

Regulation of µ and δ opioid receptor functions: involvement of cyclin-dependent kinase 5.

BACKGROUND AND PURPOSE: Phosphorylation of δ opioid receptors (DOP receptors) by cyclin-dependent kinase 5 (CDK5) was shown to regulate the trafficking of this receptor. Therefore, we aimed to determine the role of CDK5 in regulating DOP receptors in rats treated with morphine or with complete Freund's adjuvant (CFA). As µ (MOP) and DOP receptors are known to be co-regulated, we also sought to determine if CDK5-mediated regulation of DOP receptors also affects MOP receptor functions. EXPERIMENTAL APPROACH: The role of CDK5 in regulating opioid receptors in CFA- and morphine-treated rats was studied using roscovitine as a CDK inhibitor and a cell-penetrant peptide mimicking the second intracellular loop of DOP receptors (C11-DOPri2). Opioid receptor functions were assessed in vivo in a series of behavioural experiments and correlated by measuring ERK1/2 activity in dorsal root ganglia homogenates. KEY RESULTS: Chronic roscovitine treatment reduced the antinociceptive and antihyperalgesic effects of deltorphin II (Dlt II) in morphine- and CFA-treated rats respectively. Repeated administrations of C11-DOPri2 also robustly decreased Dlt II-induced analgesia. Interestingly, DAMGO-induced analgesia was significantly increased by roscovitine and C11-DOPri2. Concomitantly, in roscovitine-treated rats the Dlt II-induced ERK1/2 activation was decreased, whereas the DAMGO-induced ERK1/2 activation was increased. An acute roscovitine treatment had no effect on Dlt II- or DAMGO-induced analgesia. CONCLUSIONS AND IMPLICATIONS: Together, our results demonstrate that CDK5 is a key player in the regulation of DOP receptors in morphine- and CFA-treated rats and that the regulation of DOP receptors by CDK5 is sufficient to modulate MOP receptor functions through an indirect process.

Intrinsic neurophysiological properties of hilar ectopic and normotopic dentate granule cells in human temporal lobe epilepsy and a rat model.

Hilar ectopic dentate granule cells (DGCs) are a salient feature of aberrant plasticity in human temporal lobe epilepsy (TLE) and most rodent models of the disease. Recent evidence from rodent TLE models suggests that hilar ectopic DGCs contribute to hyperexcitability within the epileptic hippocampal network. Here we investigate the intrinsic excitability of DGCs from humans with TLE and the rat pilocarpine TLE model with the objective of comparing the neurophysiology of hilar ectopic DGCs to their normotopic counterparts in the granule cell layer (GCL). We recorded from 36 GCL and 7 hilar DGCs from human TLE tissue. Compared with GCL DGCs, hilar DGCs in patient tissue exhibited lower action potential (AP) firing rates, more depolarized AP threshold, and differed in single AP waveform, consistent with an overall decrease in excitability. To evaluate the intrinsic neurophysiology of hilar ectopic DGCs, we made recordings from retrovirus-birthdated, adult-born DGCs 2-4 mo after pilocarpine-induced status epilepticus or sham treatment in rats. Hilar DGCs from epileptic rats exhibited higher AP firing rates than normotopic DGCs from epileptic or control animals. They also displayed more depolarized resting membrane potential and wider AP waveforms, indicating an overall increase in excitability. The contrasting findings between disease and disease model may reflect differences between the late-stage disease tissue available from human surgical specimens and the earlier disease stage examined in the rat TLE model. These data represent the first neurophysiological characterization of ectopic DGCs from human hippocampus and prospectively birthdated ectopic DGCs in a rodent TLE model.

Impact of a preventive programme on the occurrence of incidents during the transport of critically ill patients.

OBJECTIVE: Incidents related to transport of critically ill patients have been extensively reported. The objective of this study was to determine the effect of an interdisciplinary preventive programme used by all intensive care unit team members involved in patients' transport on the rate of these incidents. METHODS: A clinical quality improvement audit using a prospective pre and post intervention design was performed among medical and surgical patients hospitalised in intensive care who required intra or inter-hospital transport. RESULTS: A total of 180 transports occurred in the pre-implementation phase of the study and 187 transports in the post-implementation phase. A 20% absolute reduction of incidents was observed (57.2% vs. 37.4%, p<0.001). Statistically significant reductions were obtained for the technical problems category of incidents (25% vs. 7.5%, p<0.001) as well as the problems related to patient's mobilisation category (14.4% vs. 7.5%, p=0.05). Clinically significant trends were also observed for the clinical deterioration (24.4% vs. 17.1%, p=0.11) and undesired delay before test (23.9% vs. 17.6%, p=0.14) categories but did not reach statistical significance. CONCLUSIONS: A preventive programme applied by all care providers involved in transport of critically ill patients was associated with a reduction of incidents. The application of such a programme should be acknowledged as a standard of care considering the risks inherent to the transportation of ICU patients.

Derivation of mesenchymal stem cells from human induced pluripotent stem cells cultured on synthetic substrates.

Human-induced pluripotent stem cells (hiPSCs) may represent an ideal cell source for research and applications in regenerative medicine. However, standard culture conditions that depend on the use of undefined substrates and xenogeneic medium components represent a significant obstacle to clinical translation. Recently, we reported a defined culture system for human embryonic stem cells using a synthetic polymer coating, poly[2-(methacryloyloxy)ethyl dimethyl-(3-sulfopropyl)ammonium hydroxide] (PMEDSAH), in conjunction with xenogeneic-free culture medium. Here, we tested the hypothesis that iPSCs could be maintained in an undifferentiated state in this xeno-free culture system and subsequently be differentiated into mesenchymal stem cells (iPS-MSCs). hiPSCs were cultured on PMEDSAH and differentiated into functional MSCs, as confirmed by expression of characteristic MSC markers (CD166+, CD105+, CD90+,CD73+, CD31-, CD34-, and CD45-) and their ability to differentiate in vitro into adipogenic, chondrogenic, and osteoblastic lineages. To demonstrate the potential of iPS-MSCs to regenerate bone in vivo, the newly derived cells were induced to osteoblast differentiation for 4 days and transplanted into calvaria defects in immunocompromised mice for 8 weeks. MicroCT and histologic analyses demonstrated de novo bone formation in the calvaria defects for animals treated with iPS-MSCs but not for the control group. Moreover, positive staining for human nuclear antigen and human mitochondria monoclonal antibodies confirmed the participation of the transplanted hiPS-MSCs in the regenerated bone. These results demonstrate that hiPSCs cultured in a xeno-free system have the capability to differentiate into functional MSCs with the ability to form bone in vivo.

Thromboxane A2 modulates cisplatin-induced apoptosis through a Siva1-dependent mechanism.

Thromboxane A(2) (TXA(2)) is an important lipid mediator whose function in apoptosis is the subject of conflicting reports. Here, a yeast two-hybrid screen for proteins that interact with the C-terminus of the TXA(2) receptor (TP) identified Siva1 as a new TP-interacting protein. Contradictory evidence suggests pro- and anti-apoptotic roles for Siva1. We show that a cisplatin treatment induces TXA(2) synthesis in HeLa cells. We demonstrate that endogenous TP stimulation promotes cisplatin-induced apoptosis of HeLa cells and that such modulation requires the expression of Siva1, as evidenced by inhibiting its endogenous expression using siRNAs. We reveal that, upon stimulation of TP, degradation of Siva1 is impeded, resulting in an accumulation of the protein, which translocates from the nucleus to the cytosol. Translocation of Siva1 correlates with its reduced interaction with Mdm2 (an inhibitor of p53 signalling), as well as with its increased interaction with TRAF2 and XIAP (known to enhance pro-apoptotic signalling). Our data provide a model that reconciles the pro- and anti-apoptotic roles that were reported for Siva1 and identify a new mechanism for promoting apoptosis by G protein-coupled receptors. Our findings may have implications in the use of cyclo-oxygenase inhibitors during cisplatin chemotherapy and might provide a target to reduce cisplatin toxicity on non-cancerous tissues.

Comparison of computed tomography and routine radiography of the tympanic bullae in the diagnosis of otitis media in the calf.

BACKGROUND: Otitis media is difficult to diagnose antemortem. Case reports have described computed tomography (CT) in the diagnosis, but not all cases were confirmed. HYPOTHESIS: CT is a sensitive and specific imaging modality of the tympanic bullae and can be used as the gold standard for the diagnosis of otitis media. ANIMALS: Sixteen Holstein calves 5-7 weeks of age were included. METHODS: Prospective study. All calves were sedated with i.v. xylazine (0.05-0.15 mg/kg) for routine radiography (3 views) and CT of the tympanic bullae followed by necropsy. RESULTS: Based upon necropsy findings, 10 of 16 calves were affected with otitis media, 4 unilaterally and 6 bilaterally. Imaging changes associated with otitis media included increased soft tissue opacity within the bulla, thickening of the bulla wall, enlarged bulla, and osteolysis of the bulla wall and trabeculations. The most frequent radiographic changes were lysis of trabeculations and increased soft tissue opacity, which were present in 56.3% of affected bullae. On CT, increased soft tissue opacity within the bulla was present in 93.8% of affected bullae. Sensitivity of radiography and CT was 68.8 and 93.8% and specificity was 50 and 100%, respectively. The κ value between radiography and CT with necropsy diagnosis was 0.19 for radiography, indicating poor agreement, and 0.94 for CT, indicating excellent agreement. CONCLUSION: CT is more specific, more sensitive, and easier to interpret than radiography and can be used as the gold standard in the diagnosis of otitis media in the calf.

Self-administered prescriptions of oral steroids and antibiotics in chronic obstructive pulmonary disease: are we doing more harm than good?

Patients with chronic obstructive pulmonary disease (COPD) are often given a prescription for a short course of oral steroids and antibiotics for self-administration during an acute exacerbation. The main objective of this study was to determine the impact of such prescriptions on medical care utilization, and steroids and antibiotics intake. This retrospective cohort study included patients with moderate to severe COPD participating in a self-management programme. We compared the number of unplanned medical visits (including hospitalizations) and the utilization of systemic steroids (number of short courses, number of days on treatment) and antibiotics (number of treatments) over a period of six months following registration to the programme in patients who received such a prescription and those who did not. Data were collected from hospital and community pharmacy files. A total of 89 patients were included; 46 received a self-administered prescription. During the study period, we found no difference between the two groups in the number of unplanned medical visits. However, we observed small but significant differences in the number of short courses of Prednisone (P = 0.018) and antibiotics (P = 0.006). This translated in an important difference in the number of days on steroids over the same period (;Prescription' group: 26; controls: 8; P = 0.005). Self-administered prescriptions may increase steroids and antibiotics utilization in patients with moderate to severe COPD, without reducing the number of unplanned medical visits.

Evaluation of the contribution of cyclooxygenase 1 and cyclooxygenase 2 to the production of PGE2 and PGF2 alpha in epithelial cells from bovine endometrium.

In ruminants, the production of prostaglandins by the endometrium is critical for recognition of pregnancy. In the absence of an embryonic signal, luteolytic pulses of PGF(2 alpha) are released by the uterus. In contrast, the presence of a viable conceptus reduces the production of PGF(2 alpha) relative to PGE(2) and prevents luteolysis through the release of trophoblastic interferon (IFN-tau). Initially, it was thought that epithelial and stromal endometrial cells were specialized in the production of a single type of prostaglandin. However, purified cell populations of both types of cell can produce PGF(2 alpha) and PGE(2); therefore, selective production of PGF(2 alpha) and PGE(2) must be regulated within each type of cell. Two distinct prostaglandin synthases, cyclooxygenase 1 and cyclooxygenase 2, are involved in prostaglandin production and each may catalyse the production of a different prostaglandin. This possibility was investigated in cultured epithelial cells from bovine endometrium. Cells were treated with oxytocin or arachidonic acid, and expression of cyclooxygenase 1 and cyclooxygenase 2 proteins was monitored over time and correlated with prostaglandin accumulation. Cells were also treated with increasing doses of inhibitors of cyclooxygenase 1 or cyclooxygenase 2 (non-steroidal anti-inflammatory drugs; NSAIDs) with or without arachidonic acid or oxytocin: flurbiprofen (0-50 micromol l(-1)) was used as a non-selective inhibitor; valeryl salicylate (0-500 micromol l(-1)) was used as a cyclooxygenase 1 inhibitor and NS-398 (0-1 micromol l(-1)) was used as a cyclooxygenase 2 inhibitor. After stimulation with arachidonic acid or oxytocin, prostaglandin production and expression of cyclooxygenase 2 protein were increased. All inhibitors were able to block basal and stimulated prostaglandin production. These results indicate that in endometrium most, if not all, prostaglandin production is probably processed through cyclooxygenase 2.

The immunophenotype of blood and cerebrospinal fluid mononuclear cells in dogs.

Inflammatory neurologic diseases are common in dogs, but establishing a definitive diagnosis often is difficult. Nucleated cell number and type in cerebrospinal fluid (CSF) rarely are suggestive of an etiologic agent. We speculated that CSF leukocyte immunophenotyping would be a useful adjunct in the investigation of canine inflammatory neurologic diseases by yielding more specific etiologic information. The goals of this study were to establish the feasibility of flow cytometric evaluation of individual canine CSF samples and to identify the cell distribution in healthy dogs. The mononuclear cell populations of paired blood and CSF samples from 23 healthy dogs were characterized by labeling of cells with antibodies against CD4, CD8alpha, CD21, and CD14 molecules and by flow cytometric analysis of their expression. The mean proportion of CD4+ and CD21+ cells was significantly higher in blood than in the CSF (P < .002 and P < .001, respectively). In contrast, the mean proportion of CD14+ and CD8a+ cells was not significantly different between blood and CSF (P = .5 and p = .9, respectively). These findings demonstrate differences in the distribution and function of mononuclear cells in the circulating venous and subarachnoid compartments in the dog.

Analysis of antibody gene rearrangement, usage, and specificity in chronic focal encephalitis.

BACKGROUND: Autoantibodies have been implicated in the development of chronic focal encephalitis (CFE) or Rasmussen's disease, a progressive and intractable form of epilepsy characterized by uncontrollable unilateral focal seizures, brain atrophy, and inflammation. OBJECTIVE: To investigate the origin and characteristics of the B cell population that trigger or sustain brain inflammation in patients with CFE. METHODS: The authors used immunoglobulin (Ig) complementary determining region 3 (CDR3)-size spectratyping and DNA sequencing to examine the rearranged IgG heavy chain (IgGH) transcript repertoire in resected brain samples from four patients with CFE. They also performed Western blotting on human and rat brain homogenates and immunostaining on a human neuronal cell line to test the reactivity of sera from patients with CFE. RESULTS: The authors observed substantial perturbations from the normal, unstimulated repertoire of immunoglobulin genes. Sequencing of randomly selected clones confirmed the restricted profile and provided evidence for somatic mutation patterns characteristic of antigen-specific stimulation. They also observed IgGVH-CDR3 sequence diversity among patients. When sera were assayed from patients with CFE for specificity against rat and human brain homogenates, heterogeneous reactivity patterns were detected among patients. Immunostaining of postmitotic human neuronal cells demonstrated reactivity of some patients' sera against neural antigens. CONCLUSIONS: These findings support an important role for clonally expanded B lymphocytes in some forms of epilepsy, but also indicate a wide spectrum of reactivity characteristic of antigenic heterogeneity.

Modification of a commercial electrospray nebulizer for operation in a liquid chromatography/mass spectrometry system at flow rates in the low microL/min range.

A simple and inexpensive approach to convert the electrospray nebulizer of a commercial liquid chromatography/mass spectrometry (LC/MS) system (HP 1100) to accommodate lower flow rates has been proposed and evaluated. This modification consists of simply replacing the nebulizer needle by a commercially available stainless steel needle with a smaller internal diameter. Experiments were conducted in order to optimize operational parameters. Using two different internal diameter needle sizes, flow rates ranging from 1 to 250 microL/min could be accommodated. The modification presented allows an extension of the range of compatible flow rates without major modification of the standard design of the interface.

Differential regulation of basic helix-loop-helix mRNAs in the dentate gyrus following status epilepticus.

In various chemoconvulsant models of human temporal lobe epilepsy, the induction of epileptogenesis by a prolonged period of continuous seizure activity is accompanied by significant changes in hippocampal structure. These changes include an increase in neurogenesis within the proliferative subgranular zone (SGZ) of the dentate gyrus and induction of mossy fiber sprouting in mature dentate granule cells. As dentate granule cell neurogenesis and axon outgrowth are also hallmarks of hippocampal development, we hypothesized that molecules involved in normal development may also play a role in similar changes associated with epileptogenesis. To begin to test this hypothesis, we have analyzed the expression patterns of multiple members of the basic helix-loop-helix (bHLH) family of transcription factors in both normal and epileptic adult rats. bHLH protein expression has been found recently in dentate granule cells at specific developmental stages, and analysis of developmental models suggests specific neural differentiation functions for these molecules. We show that mRNA expression of all seven bHLH family members examined in this study, as well as the divergent homeobox protein Prox1, is present in the adult. Patterns of expression varied considerably between family members, ranging from the limited expression of Mash1 in the neurogenic SGZ of the dentate gyrus to the scattered, widespread profile of Hes5 throughout the dentate gyrus and the hippocampus proper. Moreover, these varied profiles of expression were differentially regulated following status epilepticus, with some increasing (Mash1, Id2), some falling (Hes5, Prox1), and others remaining mostly unchanged (NeuroD/BETA2, NeuroD2/NDRF, Id3, Rath2/Nex1). While the function of these molecules in the adult brain remains to be characterized, our findings support the idea that molecules controlling cell-fate decisions in the developing dentate gyrus are also operative during seizure-induced neurogenesis and plasticity.

Evaluation of two different educational interventions for adult patients consulting with an acute asthma exacerbation.

Asthma education decreases the number of emergency visits in specific subgroups of patients with asthma. However, it remains unknown whether this improvement is related only to the use of an action plan alone or to other components of the educational intervention. A total of 126 patients consulting urgently for an acute asthma exacerbation were recruited; 98 completed the study. The first 45 patients were assigned to Group C (control; usual treatment). Thereafter, patients were randomized to either Group LE (limited education; teaching of the inhaler technique plus self- action plan given by the on call physician) or Group SE (same as group LE plus a structured educational program emphasizing self-capacity to manage asthma exacerbations). At baseline, there was no difference between groups in asthma morbidity, medication needs, or pulmonary function. After 12 mo, only Group SE showed a significant improvement in knowledge, willingness to adjust medications, quality of life scores, and peak expiratory flows. In the last 6 mo, the number of unscheduled medical visits for asthma was significantly lower in Group SE in comparison with groups C and LE (p = 0.03). The number (%) of patients with unscheduled medical visits also decreased significantly in Group SE compared with Groups C and LE (p = 0.02). We conclude that a structured educational intervention emphasizing self-management improves patient outcomes significantly more than a limited intervention or conventional treatment.

Role of the differentially spliced carboxyl terminus in thromboxane A2 receptor trafficking: identification of a distinct motif for tonic internalization.

The thromboxane A(2) receptor (TP) is a G protein-coupled receptor that is expressed as two alternatively spliced isoforms, alpha (343 residues) and beta (407 residues) that share the first 328 residues. We have previously shown that TPbeta, but not TPalpha, undergoes agonist-induced internalization in a dynamin-, GRK-, and arrestin-dependent manner. In the present report, we demonstrate that TPbeta, but not TPalpha, also undergoes tonic internalization. Tonic internalization of TPbeta was temperature- and dynamin-dependent and was inhibited by sucrose and NH(4)Cl treatment but unaffected by wild-type or dominant-negative GRKs or arrestins. Truncation and site-directed mutagenesis revealed that a YX(3)phi motif (where X is any residue and phi is a bulky hydrophobic residue) found in the proximal portion of the carboxyl-terminal tail of TPbeta was critical for tonic internalization but had no role in agonist-induced internalization. Interestingly, introduction of either a YX(2)phi or YX(3)phi motif in the carboxyl-terminal tail of TPalpha induced tonic internalization of this receptor. Additional analysis revealed that tonically internalized TPbeta undergoes recycling back to the cell surface suggesting that tonic internalization may play a role in maintaining an intracellular pool of TPbeta. Our data demonstrate the presence of distinct signals for tonic and agonist-induced internalization of TPbeta and represent the first report of a YX(3)phi motif involved in tonic internalization of a cell surface receptor.