SEARCH · PubMed Health

Results for “Microinjections”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources

PubMed1991

Dopamine and conditioned reinforcement. II. Contrasting effects of amphetamine microinjection into the nucleus accumbens with peptide microinjection into the ventral tegmental area.

It has been shown that infusion of certain neuropeptides into the ventral tegmental area (VTA) results in increased motor activity and enhanced dopamine turnover in the nucleus accumbens. In the present experiments, substance P (SP), neurotensin (NT), d-ala-metenkephalin (DALA) and morphine sulfate (MS) were injected bilaterally into the VTA and their effects on conditioned reinforcement were assessed. These effects were compared with infusion of amphetamine into the nucleus accumbens, which has previously been shown to strongly enhance responding for conditioned reinforcers. For these experiments, hungry rats were trained to associate a compound stimulus (light and click) with the presentation of food. In the test phase, responding on one lever (CR lever) resulted in the presentation of the stimulus but no food. Responding on the other (NCR lever) had no consequences. Different groups of animals received microinjections (0.5 microliter, bilaterally) of SP (0, 0.03, 0.3, 3.0 micrograms), NT (0, 0.025, 0.25, 0.5 microgram), DALA (0, 0.01, 0.1, 1.0 microgram) or morphine (0, 0.025, 0.25, 2.5 micrograms) into the VTA. SP infusion into the VTA resulted in a small increase in responding which was not selective for the CR lever. NT, DALA and morphine had no effect on responding for conditioned reward. In contrast, amphetamine (0, 0.2, 2.0, 20 micrograms) injected into the nucleus accumbens markedly enhanced responding for conditioned reward. These findings suggest that stimulation of the mesolimbic system at the level of the DA cell bodies, which induces a small increase in DA turnover, is not sufficient to potentiate responding for conditioned reward. On the other hand, an important requirement for potentiation may be excessive release of dopamine in the nucleus accumbens.

Animals↗

PubMed1992

Neuroethological evaluation of audiogenic seizures and audiogenic-like seizures induced by microinjection of bicuculline into the inferior colliculus. II. Effects of nigral clobazam microinjections.

Male Wistar rats were classified as susceptible (S) and resistant (R) to audiogenic seizures (AS) by evaluation of their response to high-intensity sound stimulation (110.3 dB). R rats injected with bicuculline into the inferior colliculus (IC) preferentially displayed audiogenic-like seizures with gyri, jumping and atonic falling, without important tonic-clonic components but with postictal contralateral asymmetry and hyperreactivity. These audiogenic-like seizures were blocked by clobazam microinjection into the substantia nigra (SN) and partially modified by SN vehicle injection. Injection of vehicle or clobazam into the SN of susceptible rats (S) did not modify the occurrence of AS. This may suggest the participation of GABAergic regulation in the development of audiogenic-like seizures in R rats and a defect in GABAergic neurotransmission in S rats.

Acoustic Stimulation↗

PubMed1975

Protein migration into nuclei. II. Frog oocyte nuclei accumulate a class of microinjected oocyte nuclear proteins and exclude a class of microinjected oocyte cytoplasmic proteins.

Nuclear contents or cytoplasm from Xenopus oocytes labeled with (35-S)methionine or (3-H)proline (donor oocytes) were reinjected into unlabeled oocytes (recipient oocytes). The radioactivity injected as nuclear contents was found to enter and accumulate in the recipient oocyte nucleus. In contrast, the radioactivity injected as cytoplasm was found to enter but not to accumulate in the recipient oocyte nucleus. Sodium dodecyl sulfate (SDS) gel electrophoresis of the nucleus and cytoplasm of donor oocytes revealed the existence of three classes of labeled proteins in these oocytes: those proteins found predominantly in the nucleus (N proteins), those found predominantly in the cytoplasm (C proteins), and those found in both the nucleus and cytoplasm at similar concentrations (B proteins). SDS gel electrophoresis of the nucleus and cytoplasm of recipient oocytes showed that N proteins entered and accumulated in the nucleus but that B proteins partitioned about equally between the nucleus and cytoplasm. A similar analysis of oocytes injected with labeled cytoplasm showed that C proteins did not enter the nucleus but again B proteins partitioned about equally between the nucleus and cytoplasm.

Animals↗

PubMed1992

Additive effects of dopamine and 8-OH-DPAT microinjected into the nucleus ambiguus in eliciting vagal bradycardia in rats.

The effects of combined microinjection into the nucleus ambiguus (NA) of dopamine (DA) and the 5-hydroxytryptamine-1A (5HT-1A) receptor agonist 8-hydroxy-2-[di-n-propylamino]tetralin (8-OH-DPAT) on arterial pressure (AP) and heart rate (HR) were studied in 24 urethane-anaesthetized, artificially ventilated spinal (C1) rats. Sites from which bradycardia was elicited by microinjection of L-glutamate (GLU) were selected for microinjection of DA and 8-OH-DPAT into the NA. Microinjections of 8-OH-DPAT (60-300 pmol in 10 nl) elicited a dose-dependent bradycardia; the HR responses elicited by microinjection of 300 pmol of 8-OH-DPAT were significantly larger (-34.0 +/- 3.0) than responses elicited by the threshold dose of 60 pmol (-3.1 +/- 0.1 bpm). The onset latency of the HR responses elicited by the 300 pmol of 8-OH-DPAT was 6.5 +/- 0.5 s and the peak was reached in 40.0 +/- 8.0 s. The duration of these responses was 615 +/- 3.5 s. Microinjection of DA (1 nmol in 10 nl) within 3-5 s from the time of 8-OH-DPAT microinjection (60 pmol in 2 nl) into the NA at sites previously shown to elicit decreases in HR following microinjection of GLU, produced significant additive effects in eliciting bradycardia (20.4 +/- 2.9 bpm) when compared with decreases in HR elicited by microinjection of DA (11.1 +/- 1.8 bpm) or of 8-OH-DPAT (3.1 +/- 0.1 bpm) alone. There were no changes in AP after microinjections of DA, 8-OH-DPAT or of these two substances combined.(ABSTRACT TRUNCATED AT 250 WORDS)

8-Hydroxy-2-(di-n-propylamino)tetralin↗

PubMed1991

Microinjection of glycine into the nucleus ambiguus elicits tachycardia in spinal rats.

In 30 male Wistar spinal (C1) rats, anaesthetized with urethane and artificially ventilated, experiments were done to study the effect on heart rate (HR) and arterial pressure (AP) of microinjection of the inhibitory amino acid glycine (Gly) into the nucleus ambiguus (NA). L-Glutamate (Glu; 1.5 nmol) was microinjected into the region of the right NA to search for sites from which decreases in AP and HR could be elicited. The decreases in HR were found to be 73.1 +/- 7.0 bpm (n = 30). No changes in AP were observed. Microinjection of Gly (1 M; 2-20 nmol in 2-20 nl; n = 12) elicited a dose dependent increase in HR with no changes in AP. Microinjection of Gly 1-2 min before microinjection of Glu in 7 sites reduced significantly (P less than 0.05) the decrease in HR elicited by Glu from 87.0 +/- 27.3 bpm to 17.7 +/- 7.2 bpm. Increases in HR elicited by Gly in the right NA of another 12 rats were not affected significantly by prior microinjection of the Gly antagonist strychnine hydrochloride (30-90 pmol in 10-30 nl in one group of animals, n = 6; and 2.5 nmol in 50 nl in another group, n = 6). In addition, to determine whether the effects of Gly were caused by actions on N-methyl-D-aspartate (NMDA) receptors, kynurenic acid (KYN; 4.5 nmol in 30 nl) was microinjected into the right NA of 6 rats prior to microinjection of Gly. KYN failed to block the response to Gly microinjection and instead potentiated the HR increase elicited by Gly.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

The incorporation and dispersion of cells and latex beads on microinjection into the amniotic cavity of the mouse embryo at the early-somite stage.

The ability of cells and latex beads to become incorporated into the cranial region of embryos after microinjection into the amniotic cavity was studied. Premigratory neural crest cells isolated from the lateral margins of the neuroepithelium, 3T3 fibroblast cells or H35 hepatoma cells were labelled with WGA-gold conjugates, and were then microinjected into the amniotic cavity of embryos with two to three somites in vitro. Latex beads were similarly microinjected into different groups of embryos. Incorporation of injected cells or latex beads was found in the neural crest of the midbrain and the hindbrain of 5-20% of the recipients 4 h after microinjection. At 6 and 12 h, increasingly more embryos (20-77%) were observed with labelled cells or latex beads in the crest region. While hepatoma cells and latex beads were restricted to the crest region, injected neural crest cells and fibroblasts were also found in the lateral mesenchyme, bounded laterally by the surface ectoderm and medially by the closing neural tube. By 24 h after microinjection, the injected cells or latex beads were found in 50-80% of the recipients. Neural crest cells and fibroblasts, which showed similar patterns of distribution in the embryos, were located on the dorsal aspect of the neural tube, the lateral mesenchyme, the pharyngeal arches and the regions for ganglia. Hepatoma cells and latex beads were limited to the dorsal regions of the neural tube. When microinjection was carried out in embryos with seven to eight somites, incorporation of cells or latex beads was found in 44-75% of embryos, but no dispersion of the incorporated cells or latex beads into the mesenchyme was found 24 h after microinjection. Incorporation and dispersion of cells and latex beads were not observed when embryos with 18-20 somites were used as recipients. The present study showed that neural crest or fibroblast cells when injected into the amniotic cavity could be incorporated into the neural crest, and then undergo migration along the neural crest pathways, whereas hepatoma cells and latex beads could only be incorporated. The incorporation and migration of the exogenous tissues are related to the formation and the accessibility of the neural crest in the recipients.

3T3 Cells↗

PubMed1991

Cardiovascular responses to combined microinjection of substance P and acetylcholine in the intermediolateral nucleus of the rat.

As microinjection of either substance P (SP) or acetylcholine (ACh) into the right intermediolateral cell nucleus (IML) at the T2 level elicits increases in heart rate (HR) in the anesthetized rat, we investigated the possibility of a synergistic effect on HR and arterial pressure (AP) of ACh and SP microinjected in this nucleus. Moreover, we studied the effect on HR and AP of microinjection of either ACh or SP into the IML combined with activation of cardiovascular neurons in the ipsilateral rostral ventrolateral medulla (RVLM) by microinjection of glutamate (Glu). Male Wistar rats (n = 16) were anesthetized with urethane (1.4 g/kg i.p.), artificially ventilated, and the dorsal medulla and spinal cord (T1-T3) were exposed. Micropipettes containing SP and ACh were positioned in the right IML at the T2 level. Microinjection of threshold amounts of ACh (5 x 10(-2) M, 2-10 nl) and SP (3 x 10(-6) M, 2-10 nl) that caused small or no changes in HR or AP (less than 10 bpm or mmHg) elicited statistically significant synergistic increases in HR (22.9 +/- 3.3 bpm) but no changes in AP. Threshold microinjections of Glu (0.18 M, 2-10 nl) into the right RVLM combined with microinjections of threshold amounts of SP or ACh into the ipsilateral IML elicited significant synergistic increases in HR of 13.1 +/- 1.9 bpm and 10.6 +/- 1.9 bpm and in AP of 9.7 +/- 1.9 mmHg and 10.8 +/- 1.7 mmHg, respectively. These results indicate that SP and ACh interact to influence cardioacceleratory spinal preganglionic neurons (SPN) and interact with the transmitter released in the IML by RVLM stimulation to elicit increases in HR.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗

PubMed1992

Microinjection of antibodies and expression vectors into living myocardial cells. Development of a novel approach to identify candidate genes that regulate cardiac growth and hypertrophy.

BACKGROUND: Microinjection approaches in the cardiac cell context have allowed delivery of various calcium dyes and monitoring of short-term physiological responses. However, unlike other cell types, it has proved difficult to microinject myocardial cells without the concomitant loss of long-term cell viability. METHODS AND RESULTS: An analysis of experimental variables was conducted to adapt microinjection techniques to the neonatal rat ventricular cell context. Among the variables optimized were the selection of culture dishes, plating substrate, microinjection parameters, and a variety of maneuvers to inhibit myocyte hypercontracture, injury, and consequent death after micropuncture. With the modified technique, the percentage of injected cells that maintained long-term viability (48 hours) increased from less than 1% to 30%. Similarly, an increased efficiency of gene transfer and expression (measured as the percentage of injected cells that express the delivered gene) was obtained after either cytoplasmic or nuclear injection of a beta-galactosidase expression vector into cardiac myocytes. Microinjection of marker immunoglobulin G does not interfere with the induction of the hypertrophic response or the expression of a coinjected atrial natriuretic factor promoter-luciferase reporter fusion gene construct. CONCLUSIONS: To the best of our knowledge, this study provides the first description of the efficient microinjection of neonatal cardiac muscle cells with maintenance of long-term cell viability. The microinjection technique is now a viable approach to examine cause-and-effect relations between specific gene products and any defined feature or response of cardiac myocytes that can be assayed at a single-cell level.

Animals↗

PubMed1992

Fate of microinjected genes in preimplantation mouse embryos.

The state of genes microinjected into mouse embryos was followed from the one-cell to the blastocyst stage using the polymerase chain reaction (PCR). Microinjected DNA was detected in all one-, two-, and four-cell injected embryos and in 44% of morula and 26% of blastocysts. Head-to-tail ligation of microinjected genes, a common feature of stably integrated transgene arrays, was detected in all embryos after injection of microinjected genes and occurred irrespective of the structure at the ends of the injected genes. Sensitivity of microinjected DNA to a methylation-dependent restriction endonuclease Dpn I was lost in all embryos by the two-cell stage (24 hr), indicating a change in DNA methylation, independent of transgene integration. Dissociation of blastomeres prior to compaction revealed a mosaic distribution of the microinjected DNA within the embryo and supports the notion that injected genes form a limited number of arrays, which segregate independently until they integrate into the genome or are degraded.

Animals↗

PubMed1991

Enkephalins, substance P and acetylcholine microinjected into the nucleus ambiguus elicit vagal bradycardia in rats.

Little is known about putative transmitters in the nucleus ambiguus (NA) mediating parasympathetic control of the heart, although Met-enkephalin (m-ENK), Leu-enkephalin (l-ENK), substance P (SP) and acetylcholine (Ach) have been detected in the cell bodies and fibers of this nucleus. The effects of these substances on arterial pressure (AP) and heart rate (HR) were studied by microinjecting them (4-20 nl) into the NA. Experiments were done in 26 spinal (high cervical) rats that were anesthetized with urethane and artificially ventilated. L-Glutamate (GLU) was microinjected into the right NA to identify the location of cell bodies from which decreases in HR and AP could be elicited. m-ENK, l-ENK, SP or Ach was then microinjected into these sites. Microinjection of 1 nmol of GLU elicited significant decreases in HR (-72.2 +/- 9.7 bpm, n = 15) which were not accompanied by significant decreases in mean AP. Microinjection of m-ENK (15-200 pmol; n = 7), l-ENK (15-200 pmol; n = 6), SP (0.9-15 pmol; n = 7) and Ach (2.0-20 pmol; n = 7) into the NA decreased HR in a dose-dependent manner but did not affect AP. The magnitudes of HR responses to m-ENK, l-ENK, SP and Ach were smaller but of longer duration than the changes in HR to microinjection of GLU. These results suggest a physiological role for GLU, enkephalins, SP and Ach in the vagal control of HR mediated by the NA.

Acetylcholine↗

PubMed1992

Cardiovascular effects of microinjections of quipazine into nuclei of the medulla oblongata in anaesthetized cats: comparison with L-glutamate.

Unilateral microinjections of quipazine (0.9 micrograms in 50 nl) into the subretrofacial nucleus produced hypertension and a slight tachycardia associated with an increase in renal sympathetic nerve activity. Microinjections of quipazine lateral, caudal or rostral to this nucleus failed to alter blood pressure and heart rate. Similarly, microinjections of l-glutamate (3 nmol in 15 nl) into the subretrofacial nucleus elicited hypertension, tachycardia and renal sympatho-excitation. The magnitude of the pressor response to quipazine was smaller than the response elicited by l-glutamate but its duration was longer. Microinjections of quipazine into the lateral tegmental field at l-glutamate hypertensive sites failed to alter arterial blood pressure and heart rate. In contrast, microinjections of quipazine into the caudal ventrolateral medulla or into the nucleus tractus solitarii produced hypotension and sympatho-inhibition. These effects were prevented by microinjections of the 5-HT2 receptor antagonists, LY 53857 or BW 501C. The present results indicate that stimulation of 5-HT2 receptors of the subretrofacial nucleus produces hypertension and sympatho-excitation whereas stimulation of 5-HT2 receptors in the caudal ventrolateral medulla and in the nucleus tractus solitarii produces hypotension and sympatho-inhibition.

Anesthesia↗

PubMed1992

Subzonal sperm microinjection in cases of severe male factor infertility and repeated in vitro fertilization failure.

OBJECTIVE: To examine (1) fertilization rates obtained with subzonal sperm microinjection when different numbers of sperm are injected into the perivitelline space; (2) when subzonal sperm microinjection is combined with dilute insemination; and (3) the association of semen quality characteristics with fertilization. DESIGN: Subzonal sperm microinjection and subzonal sperm microinjection combined with dilute insemination was performed in 109 and 41 cycles on patients in two clinical trials in Melbourne, Australia, and Bologna, Italy, respectively. PATIENT PARTICIPANTS: Couples who have experienced repeated in vitro fertilization failure or in whom the husband has severe male factor infertility. PRIMARY OUTCOME MEASURES: The number of oocytes fertilized after injection of different numbers of sperm into the perivitelline space, the number of patients transferred, and pregnancy outcome. RESULTS: The injection of multiple numbers of sperm into the perivitelline space failed to improve monospermic fertilization rates but caused an increase in polyspermic fertilization. In patients with initial semen parameters exhibiting greater than 50% motility or greater than 50% normal morphology fertilization rates were improved when subzonal sperm microinjection-treated eggs were incubated in a dilute insemination medium. Six pregnancies were obtained, two of which have progressed to term. CONCLUSIONS: When applied to male factor patients, the subzonal sperm microinjection technique results in a 14% to 15% fertilization rate. However, of the 102 embryos transferred only three (2.9%) fetal heart beats were obtained.

Female↗

PubMed1992

Cell cycle effects of microinjected antisense oligodeoxynucleotides to p34cdc2 kinase.

In this study the effect of antisense oligomers targeted against the mRNA transcripts of p34cdc2 kinase on G1 progression into S-phase was examined. For this purpose, antisense, sense, or nonsense oligomers were introduced directly into the cytoplasm of T98G cells grown in monolayer cultures by glass-capillary microinjection. The microinjection of antisense oligomers (but not sense or nonsense oligomers) into growth-arrested cells before serum stimulation inhibited G1 progression into S-phase. This inhibition was correlated with a reduction in the steady-state levels of nuclear p34cdc2 protein. Microinjection of antisense oligomers into cells at 2 and 6 hours after serum stimulation also resulted in a marked inhibition in the ability of cells to enter S-phase. The inhibitory effect decreased when cells were microinjected at 12 hours after serum stimulation. When cells were microinjected at 18 and 24 hours after serum stimulation, only a slight inhibition was observed. As the antisense oligomers were introduced directly into the cytoplasm of cells at each of the time points examined, the observed differences in the inhibitory effects of the antisense oligomers at later times after serum stimulation cannot be explained by differences in uptake. An alternative explanation is that after a certain threshold level of nuclear p34cdc2 protein is reached in late G1 phase; no further increase is necessary, because the cells become committed to enter S-phase. In yeast, p34cdc2 appears to play an important role in the G1/S-phase transition at a control point in late G1 phase called START (reviewed by Lewin). In mammalian cells a control point that could be equivalent to START is the "restriction point" which is defined as the time after which inhibition of protein synthesis fails to block entry into S-phase (reviewed by Pardee). The effects observed with antisense oligomers to p34cdc2 kinase are strikingly similar to what is observed when low concentrations of the drug cycloheximide are added to these cells at different times after serum stimulation; entry into S-phase is significantly inhibited when cycloheximide is added up to 12 hours postimulation. Thus, the results reported in this study are in agreement with the idea that p34cdc2 kinase plays a role in the G1/S phase transition in mammalian cells. Finally, introduction of antisense oligomers directly into the cytoplasm of cells grown in monolayer cultures by glass-capillary microinjection appears to be a viable alternative to simply adding the oligomers to the culture medium.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗

PubMed1990

Microinjection of substance P and ACh into rat intermediolateral nucleus elicits cardiovascular responses.

The effects of substance P (SP) or acetylcholine (ACh) microinjected into the intermediolateral nucleus of the spinal cord (IMLn) on arterial pressure (AP) and heart rate (HR) were investigated in 22 male Wistar rats under urethan and artificial ventilation. L-Glutamate (Glu) was microinjected into the IMLn between C7 and T4 to locate cardiovascular sites. Micropipettes containing Glu were stereotaxically positioned in 82 histologically verified sites in the IMLn between C7 and T4 on both the right and left sides. Microinjection of 4-10 nl of 0.18 M Glu in 30 of 39 explored sites at the T2 level elicited significant increases in HR (+24.2 +/- 3.1 beats/min). These changes were accompanied by significant increases in mean AP (+11.4 +/- 1.2 mmHg) at the T2 level (32/47 sites). Microinjection of 4-10 nl of SP (3 X 10(-7) to 3 X 10(-4) M) or ACh (0.005-0.5 M) in the right IMLn at the T2 level elicited increases in HR but did not affect AP. The duration of the responses to SP or ACh was significantly longer than the duration of the responses to Glu. The responses to ACh could be blocked by prior microinjection of 5 X 10(-2) M atropine. No responses were ever obtained in the left IMLn by microinjection of Glu, SP, or ACh. These results support the hypothesis that Glu, ACh, and SP mediate sympathoexcitation in the IMLn of the rat and that these excitatory responses have different temporal patterns.

Acetylcholine↗

PubMed1992

In vitro development of zygotes from prepubertal gilts after microinjection of DNA.

The effect of pronuclear microinjection of DNA and culture in excised mouse oviducts on the development of porcine zygotes was assessed in this study. Precocious ovulation was induced in prepubertal gilts with pregnant mare's serum gonadotrophin and hCG. Zygotes received either pronuclear microinjection of buffer alone, buffer containing a DNA construct, or no microinjection. Zygotes were cultured in vitro in either modified Krebs-Ringer bicarbonate medium (KRB) for 144 h or in mouse oviduct (MO) explant culture with KRB for 48, 72, 96, or 120 h. Pronuclear microinjection of DNA resulted in a lower (P less than .05) cleavage index (CI) than did buffer or no microinjection (CI 2.16 +/- .10 vs 2.80 +/- .13 and 2.93 +/- .10). The CI loss was greatest for DNA-injected zygotes at the two-cell stage of development. Coculture of zygotes in MO resulted in a higher CI (P less than .01) than did culture in KRB. Culture in MO for 72 h was the most beneficial system compared with MO for 48, 96, or 120 h (P less than .05; CI 3.25 +/- .12 vs 2.66 +/- .18, 2.79 +/- .14, and 2.40 +/- .14, respectively). Microinjection of DNA, not merely the mechanical procedure, was detrimental to early zygote development and may be the cause of low pregnancy rates.

Animals↗

PubMedDate not supplied

Expression of thermotolerance following microinjection of poly(A)RNA isolated from thermotolerant CHO cells.

Poly(A)RNA was isolated from thermotolerant cells and microinjected into recipient non-tolerant Chinese hamster ovary (CHO) cells. The injected cells expressed thermotolerance to a subsequent test heat treatment both in terms of the end-points of colony formation (cell survival) and resumption of protein synthesis after test heating (translational labelling). The magnitude of thermotolerance expression was dependent on the experimental end-point (increase up to 3.8-fold for translational labelling and approximately 2-fold for survival) and on the time between microinjection and the test heat treatment. Control experiments showed that poly(A)RNA from non-tolerant cells did not alter the heat response of microinjected cells. Proteins corresponding to the poly(A)RNA from thermotolerant cells were analysed by in vitro translation and by labelling of microinjected cells, followed by SDS-PAGE. In vitro translations showed high levels of transcripts for classical heat-shock proteins (HSP 70/72, 89, 110) in poly(A)RNA from thermotolerant versus control cells. However, proteins synthesized in intact cells showed no detectable differences when cells were microinjected with poly(A)RNA from thermotolerant versus control cells, or not injected at all. In principle the data show that microinjection of specific poly(A)RNA fractions can be used for defining the contribution of individual gene products to the cellular heat response.

Animals↗

PubMed1992

A comparison between in vitro fertilization and microinjection of immobilized spermatozoa from bulls producing spermatozoa with defects.

The objectives of this study were to compare the fertilization rate of bovine in vitro matured oocytes by in vitro fertilization (IVF) and by microinjection of a single spermatozoon (MI) and to relate these rates with fertility reported for these bulls in artificial breeding. Bull A (Holstein) had a nonreturn rate of 75%. Semen from this bull is routinely used in our standard IVF procedure. Bull B (Ayrshire), used regularly in artificial breeding and related to bull D, had a nonreturn rate of 69.2%. Bull C (Brown Swiss), with a chromosomal translocation and trisomy, achieved a nonreturn rate of 42%. Bull D (Ayrshire) produced nonmotile spermatozoa (SPZ) and had an abnormality described as "tail stump defect." No pregnancies sired by bull D have been reported. Oocytes were either fertilized in vitro by capacitated SPZ or by microinjection of a single immobilized SPZ into the ooplasm. SPZ were treated with 0.1 microM A23187 and used for IVF. For microinjection SPZ were cocultured for 5 h with bovine oviduct epithelial cells (BOEC) and then immobilized by freezing and thawing twice without cryoprotectant. A single batch of killed SPZ (stored at -25 degrees C) was used for all microinjections. All oocytes were cultured in Medium 199 for 22 h at 39 degrees C and subsequently fixed, stained, and examined for evidence of fertilization (i.e., female and male pronucleus formation, SPZ decondensation). Fertilization rates following IVF with semen from bulls A, B, C, and D were 80%, 54%, 1%, and 2%, and following microinjection were 39%, 22%, 21%, and 34%, respectively.

Animals↗

PubMedDate not supplied

Dynamic analysis of drug action on in vitro reconstituted thyroid follicle by microinjection of tracer molecules and videomicroscopy.

Thyroid cells isolated from the gland by trypsinization are capable in culture of reconstituting histiotypic structures, the thyroid follicles. This morphological differentiation requires the presence of the main thyroid regulator; thyrotropin. We have analyzed some structural and functional aspects of in vitro reconstituted thyroid follicles (RTF) using microinjection of fluorescent probes and videomicroscopy. This experimental approach allowed to visualize biological processes and actions of drugs, signalling factors, etc. in living cells. We describe here some examples of what can be studied with this powerful still-undervalued method. Microinjection of a cell-impermeant fluorescent probe of either high or low molecular mass into the lumen of RTF allowed to check the tightness of this compartment and therefore to analyze the control of tight junctions assembly. A small cell-impermeant probe like Lucifer Yellow microinjected into a cell was used to demonstrate and then to study the regulation of cell to cell communication via gap junctions. The presence of calcium in the lumen of RTF was detected by microinjection of a properly designed probe: Calcium Green which becomes fluorescent in the presence of the ligand. The lumen to cell transport or endocytosis of thyroglobulin, the thyroid prohormone, which is stored into the lumen of the follicles, is currently studied by microinjection of TRITC-labeled thyroglobulin. Coupled to image processing and videorecorder systems, kinetic analysis and quantitative measurements can be performed.

Animals↗